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lördag 7 september 2019

SHEARS SB. Inositolipyrofosfaatit; (InsPP) Miksi niin monta fosfaattia inositolijärjestelmällä?

https://www.ncbi.nlm.nih.gov/pubmed/25453220

2015 Jan;57:203-16. doi: 10.1016/j.jbior.2014.09.015. Epub 2014 Oct 5.

Inositol pyrophosphates: why so many phosphates?

Shears SB1.
1
Inositol Signaling Group, Laboratory of Signal Transduction, National Institute of Environmental Health Sciences, NIH, DHHS, PO Box 12233, Research Triangle Park, NC 27709, USA. Electronic address: Shears@niehs.nih.gov.

Abstract

The inositol pyrophosphates (PP-InsPs) are a specialized group of "energetic" signaling molecules found in yeasts, plants and animals. PP-InsPs boast the most crowded three dimensional phosphate arrays found in Nature; multiple phosphates and diphosphates are crammed around the six-carbon, inositol ring. Yet, phosphate esters are also a major energy currency in cells. So the synthesis of PP-InsPs, and the maintenance of their levels in the face of a high rate of ongoing turnover, all requires significant bioenergetic input. What are the particular properties of PP-InsPs that repay this investment of cellular energy? Potential answers to that question are discussed here, against the backdrop of a recent hypothesis that signaling by PP-InsPs is evolutionarily ancient. The latter idea is extended herein, with the proposal that the primordial origins of PP-InsPs is reflected in the apparent lack of isomeric specificity of certain of their actions. Nevertheless, there are other aspects of signaling by these polyphosphates that are more selective for a particular PP-InsP isomer. Consideration of the nature of both specific and non-specific effects of PP-InsPs can help rationalize why such molecules possess so many phosphates.
Published by Elsevier Ltd.

KEYWORDS:

Analogs; Cell-signaling; Diphosphoinositol polyphosphates; Inositol pyrophosphates; Kinase; Phosphorylation; Structure
PMID:
25453220
PMCID:
PMC4291286
DOI:
10.1016/j.jbior.2014.09.015
[Indexed for MEDLINE]
Free PMC Article

KOSKA ON INOSITOLIPYROFOSFAATTEJA, on myös INOSITOLIPYROFOSFATAASEJA:

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TAKESHI IJUIN (2019) artikkeli ja kuvat inositoliaineenvaihdunnan fosfataaseista mm. Syöpäsoludynamiikkaa pohditaan.

 Koska inositoliaineenvaihdunta  kiertää   vesiliukoisesta  ja rasvaliukoiseen aitioon  ja takaisin  on  fosfataaseissa niitä, jotka  vaikuttavat a)   lipidiliukoisiin fosfoinositideihin (PI, lipositolit) ja  b)  vesiliukoisiin  jonistolifosfaattilajeihin (IPx)
 Sisällysluettelossa on  seuraavia vesiliukoisia fosfataaseja mainittu:
INPP4A, Inositolipolyfosfaatti 4-fosfataasi tyyppi 1
INPP4B, Inositolipolyfosfaatti 4-fosfataasi tyyppi 2
INPP5J, Inositolipolyfosfaatti 5-fosfataasi J
INPP5K, Inositolipolyfosfatti 5-fosfataasi K
PIPP, proliinipitoinen inositolipolyfosfaattifosfataasi 
SHIP1,  SH2 domeenin sisältävä  inositolifosfaasi 1
SHIP2,  SH2 domeenin  sisältävä isositolifosfataasi  2
SKIP, lihakseen ja munuaiseen rikastunut inositol polyfosfaattifosfataasi


Rasvaliukoiseen  fosfoinositidiin vaikuttaa
PTEN , fosfataasi ja tensiinihomologi
PTPN9, PTP-MEG2 (T-soluissa polyfosfoinositideihin esim.)
Myotubulariini MTMR2 on fosfoinositidifosfataasi .

https://www.sciencedirect.com/science/article/pii/S1044579X18301159?via%3Dihub

Review
Phosphoinositide phosphatases in cancer cell dynamics—Beyond PI3K and PTEN

Under a Creative Commons license
open access
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Abstract

Phosphoinositides (PIs)  are a group of lipids that regulate intracellular signaling and subcellular biological events. The signaling by PIP3,  phosphatidylinositol-3,4,5-trisphosphate and Akt mediates the action of growth factors that are essential for cell proliferation, gene transcription, cell migration, and polarity. The hyperactivation of this signaling has been identified in different cancer cells; and, it has been implicated in oncogenic transformation and cancer cell malignancy. Recent studies have argued the role of phosphoinositides  (PIs) in cancer cell dynamics, including actin cytoskeletal rearrangement at the plasma membrane and the organization of intracellular compartments. The focus of this review is to summarize the impact of the activities of phosphoinositide (PI) phosphatases on intracellular signaling related to cancer cell dynamics and to discuss how the abnormalities in the activities of the enzymes alter the levels of phosphoinositides (PIs, lipositols) in cancer cells.

Abbreviations

ER.   endoplasmic reticulum
ERM.   ezrin, radixin, and Moesin
ESCC.  esophageal squamous cell carcinoma
FA. focal adhesion
FAK. focal adhesion kinase
FEME. fast endophilin-mediated endocytosis
GOLPH3. Golgi phosphoprotein 3
INPP4A. inositol polyphosphate 4-phosphatase type I
INPP4B. inositol polyphosphate 4-phosphatase type II
Lpd. lamellipodin
INPP5J. inositol polyphosphate 5-phosphatase J
INPP5K. inositol polyphosphate 5-phosphatase K
LOH. loss of heterogeneity
N-WASP. neural Wiskott–Aldrich syndrome protein
PDK1. phosphoinositide-dependent kinase 1
PH. plextrin homology
PI. phosphatidylinositol
PI(3)P. phosphatidylinositol-3-monophosphate
PI(4)P. phosphatidylinositol-4-monophosphate
PI(5)P. phosphatidylinositol-5-monophosphate
PI(3,4)P2. phosphatidylinositol-3,4-bisphosphate
PI(3,5)P2. phosphatidylinositol-3,5-bisphosphate
PI(4,5)P2. phosphatidylinositol-4,5-bisphosphate
PIP3. phosphatidylinositol-3,4,5-trisphosphate
PI3K. PI 3-kinase
PI4K. phosphatidylinositol 4-kinase
PIP5K. phosphatidylinositol 4-phosphate 5-kinase
PIPP. proline-rich inositol polyphosphate 5-phosphatase
PM. plasma membrane
P-Rex. phosphatidylinositol 3,4,5-trisphosphate-dependent Rac exchanger
PTEN. phosphatase and tensin homolog
PX. phox homology
SAC. suppressor of actin
SHIP1,  SH2 domain-containing inositol phosphatase 1
SHIP2,  SH2 domain-containing inositol phosphatase 2
SKIP,  skeletal muscle and kidney enriched inositol polyphosphate phosphatase
SYNJ2,  synaptojanin 2
Tks5,  tyrosine kinase substrate 5

Keywords

Cell migration
Cell polarity
Focal adhesion
Invadopodia
PI(3,4)P2
PIP3
Phosphoinositide phosphatase
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CHEN MJ (2017) .Proteiinifosfataasien genomi ja evoluutio.

https://www.ncbi.nlm.nih.gov/pubmed/28400531
2017 Apr 11;10(474). pii: eaag1796. doi: 10.1126/scisignal.aag1796.

Genomics and evolution of protein phosphatases.

Chen MJ1,2, Dixon JE3, Manning G4,2.
1
Department of Bioinformatics and Computational Biology, Genentech Inc., South San Francisco, CA 94080, USA.
2
Razavi Newman Center for Bioinformatics, Salk Institute for Biological Studies, La Jolla, CA 94027, USA.
3
Departments of Pharmacology, Cellular and Molecular Medicine, and Chemistry and Biochemistry, University of California, San Diego, La Jolla, CA 92093, USA.
4
Department of Bioinformatics and Computational Biology, Genentech Inc., South San Francisco, CA 94080, USA. manning@manninglab.org.

Tiivistelmä  Abstract

 PROTEIINIFOSFATAASIT ovat essentiellejä proteiinikinaasien opponentteja.
Yhdessä nämä entsyymit säätelevät kaikkien proteiinien fosforyloitumisen ja useimmat soluprosessit.  Jota voidaan paremmin käsittää proteiinifosforylaation  yleisosuutta, tutkijat  tekivät luettelon ihmisproteiinioen fosfatomista, joka käsittää 189  tunnettua ja oletettua ihmisen proteiinifosfataasigeeniä.  He tunnistivat myös 79 proteiinifosfataasipseudogeeniä tai retrogeeniä, joista joillakin säättaa olla jäännösfunktioita. He jäöjittivät fosfataasien  alkuperää ja diversiteettiä  rakentamalla proteiinifosfatomeja kahdeksasta muusta  eukaryosyytistä  protistista Dictyostelium  merisiiliin ( sea urchin) .  He luokittelivat  proteiinifosfataasit  kaikista yhdeksästä lajista kymmenen proteiinilaskoksen, 21 perheen ja 178 alaperheen hierarkiaan. he havaitsivat, että  yli 80% ihmisalaperheistä oli konservoituneita kautta  eläinkunnan. , mutta osoitti huomattavaaeroavuuksia  evoluutiossa, kuten ksittäisten alaperheiden  katoamisia ja  laajenemisia ja muutoksia liitedomeeneissa.  Proteiinifosfataasit osoittavat  samankaltaista  evolutionaalista dynamiikkaa kuin  proteiinikinaasit, huomattavin   katoamisin useimmissa  malliorganismeissa. Sekvenssianalyysin mukaan voidaan olettaa, että 26 ihmsen  proteiinifosfataasidomeenia on   menettänyt katalyyttisen kyvyn ja tälläionen  kyvyttömyys  on konservoituneinta  kautta ortologien.  Tällainen genominen ja evolutionaarinen näkökohta proteiinifosfataasesita antaa  kehiötä  eläinkunnan  proteiinifosforylaation yleisanalyysiin.

Protein phosphatases are the essential opposite to protein kinases; together, these enzymes regulate all protein phosphorylation and most cellular processes. To better understand the global roles of protein phosphorylation, we cataloged the human protein phosphatome, composed of 189 known and predicted human protein phosphatase genes. We also identified 79 protein phosphatase pseudogenes or retrogenes, some of which may have residual function. We traced the origin and diversity of phosphatases by building protein phosphatomes for eight other eukaryotes, from the protist Dictyostelium to the sea urchin. We classified protein phosphatases from all nine species into a hierarchy of 10 protein folds, 21 families, and 178 subfamilies. We found that >80% of the 101 human subfamilies were conserved across the animal kingdom, but show substantial differences in evolution, including losses and expansions of individual subfamilies and changes in accessory domains. Protein phosphatases show similar evolutionary dynamics to those of kinases, with substantial losses in major model organisms. Sequence analysis predicts that 26 human protein phosphatase domains are catalytically disabled and that this disability is mostly conserved across orthologs. This genomic and evolutionary perspective on protein phosphatases provides a framework for global analysis of protein phosphorylation throughout the animal kingdom.
Copyright © 2017, American Association for the Advancement of Science.
PMID:
28400531
DOI:
10.1126/scisignal.aag1796
[Indexed for MEDLINE]
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CARMAN GM (1997). DAG- pyrofosfataasi, PA fosfataasi(fosfatidihappofosfataasi) .

https://www.ncbi.nlm.nih.gov/pubmed/9370315

Phosphatidate phosphatases and diacylglycerol pyrophosphate phosphatases in Saccharomyces cerevisiae and Escherichia coli.

Carman GM1.

1
Department of Food Science, Cook College, New Jersey Agricultural Experiment Station, Rutgers University, New Brunswick 08903, USA. carman@aesop.rutgers.edu

Abstract

Phosphatidate phosphatase  (PA phosphatase) plays a major role in the synthesis of phospholipids and triacylglycerols in the yeast Saccharomyces cerevisiae. Membrane- and cytosolic-associated forms of the enzyme have been isolated and characterized. These enzymes are Mg2+-dependent and N-ethylmaleimide-sensitive. The expression of a membrane-associated form of phosphatidate phosphatase is regulated by growth phase and inositol supplementation, whereas enzyme activity is regulated by lipids, nucleotides, and by phosphorylation. Phosphatidate phosphatase is coordinately regulated with other phospholipid biosynthetic enzymes including phosphatidylserine synthase.

Diacylglycerol pyrophosphate phosphatase   is a novel enzyme of phospholipid metabolism which is present in S. cerevisiae, Escherichia coli, and mammalian cells. This enzyme possesses a phosphatidate phosphatase activity which is Mg2+-independent and N-ethylmaleimide-insensitive and is distinct from the Mg2+-dependent and N-ethylmaleimide-sensitive form of phosphatidate phosphatase.

 Genes encoding for diacylglycerol pyrophosphate phosphatase have been isolated from S. cerevisiae and E. coli. The deduced protein sequences of these genes show homology to the sequence of the mouse PAP2 (Mg2+-independent and N-ethylmaleimide-insensitive phosphatidate phosphatase) protein, especially in a novel phosphatase sequence motif. Rat liver PAP2 displays diacylglycerol pyrophosphate phosphatase activity.
PMID:
9370315
DOI:
10.1016/s0005-2760(97)00095-7
[Indexed for MEDLINE]



DAG PPi fosfataasit:

PLPP7,

https://www.ncbi.nlm.nih.gov/gene/?term=Homo+sapiens+PLPP7

Official Symbol
PLPP7provided by HGNC
Official Full Name
phospholipid phosphatase 7 (inactive)provided by HGNC
Also known as
NET39; C9orf67; PPAPDC3
Expression
Biased expression in heart (RPKM 14.9), brain (RPKM 2.9) and 9 other tissues See more
Orthologs
mouse all
Preferred Names
inactive phospholipid phosphatase 7
Names
epididymis secretory sperm binding protein
nuclear envelope transmembrane protein NET39
phosphatidic acid phosphatase type 2 domain containing 3
phosphatidic acid phosphatase type 2 domain-containing protein 3
probable lipid phosphate phosphatase PPAPDC3
 https://www.ncbi.nlm.nih.gov/pubmed/19704009/
Muscle homeostasis  

PLPP6,

  https://www.ncbi.nlm.nih.gov/gene/403313

Official Full Name
phospholipid phosphatase 6provided by HGNC
Also known as
PDP1; PSDP; PPAPDC2; bA6J24.6
Preferred Names
phospholipid phosphatase 6
Names
PPAP2 domain-containing protein 2
phosphatidic acid phosphatase type 2 domain containing 2
phosphatidic acid phosphatase type 2 domain-containing protein 2
polyisoprenoid diphosphate phosphatase type 1
presqualene diphosphate phosphatase (PSDP)

PLPP5 
Official Symbol
PLPP5provided by HGNC
Official Full Name
phospholipid phosphatase 5 provided by HGNC
https://www.ncbi.nlm.nih.gov/gene/?term=(DPPL1%3B+HTPAP%3B+PPAPDC1BM
DPPL1; HTPAP; PPAPDC1BM
Preferred Names
phospholipid phosphatase 5
Names
diacylglycerol pyrophosphate like 1
diacylglycerol pyrophosphate phosphatase-like 1
phosphatidate phosphatase PPAPDC1B
phosphatidic acid phosphatase type 2 domain containing 1B
phosphatidic acid phosphatase type 2 domain-containing protein 1B
testicular secretory protein Li 38
 PLPP4    ( expression BRAIN)

Official Symbol
PLPP4provided by HGNC
Official Full Name
phospholipid phosphatase 4provided by HGNC
Primary source
HGNC:HGNC:23531
See related
Ensembl:ENSG00000203805
Gene type
protein coding
RefSeq status
VALIDATED
Organism
Homo sapiens
Lineage
Eukaryota; Metazoa; Chordata; Craniata; Vertebrata; Euteleostomi; Mammalia; Eutheria; Euarchontoglires; Primates; Haplorrhini; Catarrhini; Hominidae; Homo
Also known as
DPPL2; PPAPDC1; PPAPDC1A
Expression
Low expression observed in reference dataset See more
Orthologs
mouse all
Preferred Names
phospholipid phosphatase 4
Names
diacylglycerol pyrophosphate like 2
diacylglycerol pyrophosphate phosphatase-like 2
phosphatidate phosphatase PPAPDC1A
phosphatidic acid phosphatase type 2 domain containing 1A
phosphatidic acid phosphatase type 2 domain-containing protein 1A

 PLPP3 , Phospholipid phosphatase 3

https://www.ncbi.nlm.nih.gov/gene/8613

Also known as
LPP3; VCIP; Dri42; PAP2B; PPAP2B
Summary
The protein encoded by this gene is a member of the phosphatidic acid phosphatase (PAP) family. PAPs convert phosphatidic acid to diacylglycerol, and function in de novo synthesis of glycerolipids as well as in receptor-activated signal transduction mediated by phospholipase D. This protein is a membrane glycoprotein localized at the cell plasma membrane. It has been shown to actively hydrolyze extracellular lysophosphatidic acid and short-chain phosphatidic acid. The expression of this gene is found to be enhanced by epidermal growth factor in Hela cells. [provided by RefSeq, Mar 2010]
Expression
Broad expression in thyroid (RPKM 205.6), placenta (RPKM 102.2) and 23 other tissues See more
Preferred Names
phospholipid phosphatase 3
Names
PAP2 beta
lipid phosphate phosphohydrolase 3
phosphatidate phosphohydrolase type 2b
phosphatidic acid phosphatase type 2B
type-2 phosphatidic acid phosphatase-beta
vascular endothelial growth factor and type I collagen inducible

We identified vascular endothelial growth factor and type I collagen inducible protein (VCIP), also known as phosphatidic acid phosphatase 2b (PAP2b), in a functional assay of angiogenesis. 

  PLPP2

 https://www.ncbi.nlm.nih.gov/gene/8612

Also known as
LPP2; PAP-2c; PAP2-g; PPAP2C

Preferred Names
phospholipid phosphatase 2
Names
PAP2-gamma
PAP2c
lipid phosphate phosphohydrolase 2, (LPP2)
phosphatidate phosphohydrolase type 2c
phosphatidic acid phosphatase 2c
phosphatidic acid phosphatase type 2C, (PPAP2C)
phosphatidic acid phosphohydrolase type 2c
type-2 phosphatidic acid phosphatase-gamma,  (PAP2-g)

The lung contains two distinct forms of phosphatidic acid phosphatase (PAP). PAP1 is a cytosolic enzyme that is activated through fatty acid-induced translocation to the endoplasmic reticulum, where it converts phosphatidic acid (PA) to diacylglycerol (DAG) for the biosynthesis of phospholipids and neutral lipids. PAP1 is Mg(2+) dependent and sulfhydryl reagent sensitive. PAP2 is a six-transmembrane-domain integral protein localized to the plasma membrane. Because PAP2 degrades sphingosine-1-phosphate (S1P) and ceramide-1-phosphate in addition to PA and lyso-PA, it has been renamed lipid phosphate phosphohydrolase (LPP). LPP is Mg(2+) independent and sulfhydryl reagent insensitive. This review describes LPP isoforms found in the lung and their location in signaling platforms (rafts/caveolae). Pulmonary LPPs likely function in the phospholipase D pathway, thereby controlling surfactant secretion. Through lowering the levels of lyso-PA and S1P, which serve as agonists for endothelial differentiation gene receptors, LPPs regulate cell division, differentiation, apoptosis, and mobility. LPP activity could also influence transdifferentiation of alveolar type II to type I cells. It is considered likely that these lipid phosphohydrolases have critical roles in lung morphogenesis and in acute lung injury and repair.

PLPP1

https://www.ncbi.nlm.nih.gov/gene/8611 

Official Symbol
PLPP1provided by HGNC
Official Full Name
phospholipid phosphatase 1provided by HGNC
Primary source
HGNC:HGNC:9228
See related
Ensembl:ENSG00000067113 MIM:607124
Gene type
protein coding
RefSeq status
REVIEWED
Organism
Homo sapiens
Lineage
Eukaryota; Metazoa; Chordata; Craniata; Vertebrata; Euteleostomi; Mammalia; Eutheria; Euarchontoglires; Primates; Haplorrhini; Catarrhini; Hominidae; Homo
Also known as
LPP1; PAP2; LLP1a; PAP-2a; PPAP2A
Summary
The protein encoded by this gene is a member of the phosphatidic acid phosphatase (PAP) family. PAPs convert phosphatidic acid (PA)  to diacylglycerol (DAG), and function in synthesis of glycerolipids and in phospholipase D-mediated signal transduction. This enzyme is an integral membrane glycoprotein that plays a role in the hydrolysis and uptake of lipids from extracellular space. Alternate splicing results in multiple transcript variants of this gene. [provided by RefSeq, May 2013]
Expression
Broad expression in prostate (RPKM 96.2), gall bladder (RPKM 74.3) and 22 other tissues See more
Orthologs
mouse all
Preferred Names
phospholipid phosphatase 1
Names
lipid phosphate phosphohydrolase 1a
phosphatidate phosphohydrolase type 2a
phosphatidic acid phosphatase 2a
phosphatidic acid phosphatase type 2A
phosphatidic acid phosphohydrolase type 2a
type-2 phosphatidic acid phosphatase alpha
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ZHAO H (2019).Pyrofosfataasien havaitseminen fluoresoivalla menetelmllä.

https://www.ncbi.nlm.nih.gov/pubmed/30572018

2019 Feb;43:29-33. doi: 10.1016/j.mcp.2018.12.003. Epub 2018 Dec 17.

A novel fluorometric method for inorganic pyrophosphatase detection based on G-quadruplex-thioflavin T.

Zhao H1, Ma C2, Chen M1.

1
School of Life Sciences, Central South University, Changsha, 410013, China.
2
School of Life Sciences, Central South University, Changsha, 410013, China. Electronic address: macb2012@csu.edu.cn.

Abstract

In this paper, we propose a fluorometric approach for the highly sensitive detection of inorganic pyrophosphatase (PPase) based on G-quadruplex-thioflavin T (ThT). In the absence of PPase, Cu2+ can coordinate with pyrophosphate (PPi) to generate a Cu2+/PPi complex. Then the G-rich sequence folds into the G-quadruplex structure, which can combine with ThT to generate a remarkable fluorescent signal. In the presence of PPase, the coordinated compound can be destroyed by the PPase catalyzed hydrolysis of PPi into inorganic phosphate (Pi). The subsequent release of Cu2+ can compete with ThT to induce a tighter G-quadruplex structure, causing the release of ThT and a sharp fluorescence decrease. Based on this mechanism, a facile and quantitative strategy for PPase detection was developed. The fluorescence intensity of the system shows a linear relationship with the PPase activities in the range of 0.5-30 U/L with a detection limit as low as 0.48 U/L. The proposed strategy for fluorescence spectrometric PPase detection is convenient, cost effective, and sensitive. This can be utilized to evaluate the inhibition effect of NaF on PPase as well as diagnose PPase-related diseases.
Copyright © 2018 Elsevier Ltd. All rights reserved.

KEYWORDS:

Fluorescence; G-quadruplex-thioflavin T; Label-free; Pyrophosphatase
PMID:
30572018
DOI:
10.1016/j.mcp.2018.12.003
[Indexed for MEDLINE]
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WANG Y (2018) Kuinka voi havaita pyrofosfataaseja? Uusi elektrokemiallinen menetelmä luotu 2017.

yksi syy miksi fosfaattien tutkimus on erittäin aktuellia;:  havaitsemistekniikka tästä  fosforin ja  energia-alueella  signalointia suorittavien proteiinien ja  mineraalien   ja fosforiyhdisteiden dynamiikasta  on  suuresti kehittynyt  vuosituhannen vaihteen jälkeen.  Esim  fytiinin  aineenvaihdunt mudoosta  vähän väliä pyrofosfaatteja samoin kuin sokeriaineenvaihdunta.  Entsyymit, jotka sitten  katalysoivat  jonien "salamannopeaita " ajallisia ja  virrallisia  vektoreita,  ovat nyt vuorossa tulla havaituiksi.

https://www.ncbi.nlm.nih.gov/pubmed/29136853

2018 Feb 1;178:491-497. doi: 10.1016/j.talanta.2017.09.069. Epub 2017 Sep 28.

Electrochemical strategy for pyrophosphatase detection Based on the peroxidase-like activity of G-quadruplex-Cu2+ DNAzyme.

Wang Y1, Wu Y2, Liu W3, Chu L3, Liao Z3, Guo W3, Liu GQ4, He X5, Wang K6.


1
Hunan Provincial Key Laboratory for Forestry Biotechnology, College of Life Science and Technology, Central South University of Forestry and Technology, 410004 Changsha, PR China; State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Biology, College of Chemistry and Chemical Engineering, Hunan University, Changsha 410082, PR China. Electronic address: bionano@163.com.
2
Hunan Provincial Key Laboratory for Forestry Biotechnology, College of Life Science and Technology, Central South University of Forestry and Technology, 410004 Changsha, PR China; State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Biology, College of Chemistry and Chemical Engineering, Hunan University, Changsha 410082, PR China.
3
Hunan Provincial Key Laboratory for Forestry Biotechnology, College of Life Science and Technology, Central South University of Forestry and Technology, 410004 Changsha, PR China.
4
Hunan Provincial Key Laboratory for Forestry Biotechnology, College of Life Science and Technology, Central South University of Forestry and Technology, 410004 Changsha, PR China. Electronic address: gaoliuedu@163.com.
5
State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Biology, College of Chemistry and Chemical Engineering, Hunan University, Changsha 410082, PR China.
6
State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Biology, College of Chemistry and Chemical Engineering, Hunan University, Changsha 410082, PR China. Electronic address: kmwang@hnu.edu.cn.

 TIIVISTELMÄ, Abstract

 Uusi yksinkertainen ja hyvin herkkä elektrokemiallinen metodi pyrofosfataasiaktiivisuuden havaitsemiseen  on kehitetty  ja se perustuu G-quadruplex-Cu2+ DNAzyymin peroksidaasinkaltaiseen aktiivisuuteen.
Kun  pyrofosfataasientsyymiä ei ole läsnä, Cu2+ voisi koordinoitua pyrofosfaatin (PPi) kanssa  muodostaen  Cu2+-PPi yhdistyksen. Mutta jos on läsnä  pyrofosfataasia (PPaasi),  entsyymi  voisi hävittää  koordinoituneen yhdisteen, koska  PPaasi katalysoi pyrofosfaatin (PPi) hydrolyysin epäorgaaniseksi fosfaatiksi Pi ja tuottaa vapaata  Cu2+ jonia, joka sitten pystyy kytkeytymään G-pitoiseen DNA:han muodostaen G-quadruplex-Cu2+DNAzyymin.   G-quadruplex-Cu2+DNAzyymin muodostuminen  immobilisoitui SPGE-pinnalla (kultaelektrodilla).  Kun käytettiin   redox-väliaineena TMB ja vetyperoksidia (H2O2 entsyymisubstraattina ,DNAzyymi katalysoi vetyperoksidin reduktion kehkeyttäen mitattavan,  kvantitatiivisen kronoamperometrisen signaalin.  Mainitun G-quadruplex-Cu2+-DNAzyymin  katalyyttinen aktiviteetti TMB-H2O2- reaktiossa oli suhteessa  pyrofosfataasin aktiivisuuteen - ja täten oli saatu  ensimmäistä kertaa  kehitettyä  yksinkertainen  ja herkkä  pyrofosfataasiaktiivisuudesta riippuva  elektrokemiallinen metodi. Järjestelmän kronoamperometrinen intensiteetti omaa eräänlaisen lineaarin suhteen  pyrofosfataasin aktiivisuuksiin rajoissa 1.0- 50.0 mU/mL.  ja havaitsemisen raja  voisi olla niinkin alhaalla kuin 0.6 mU/mL (S/N=3). Tämä esitetty metodi oli selektiivinen,  taloudellinen ja soveltuvainen,  omaa laatuaan,  ilman  monimutkaisia operaatioita   ja sitä on edelleen sovellettu  pyrofosfataasien inhibiitorien seulontaan hyvin tehokkaasti. 

A new simple and highly sensitive electrochemical method for pyrophosphatase (PPase) activity detection was developed based on the peroxidase-like activity of G-quadruplex-Cu2+ DNAzyme. In the absence of PPase, Cu2+ could coordinate with pyrophosphate (PPi) to form Cu2+-PPi compound. While in the presence of PPase, it could destroy the coordinate compound because PPase catalyzed the hydrolysis of PPi into inorganic phosphate and produced free Cu2+, which then could be coupled with G-rich DNA to form G-quadruplex-Cu2+ DNAzyme. The formation of a mimic enzyme (G-quadruplex-Cu2+ DNAzyme) was immobilized on the surface of screen-printed gold electrode (SPGE). Using 3, 3', 5, 5'-tetramethylbenzidine (TMB) as a redox mediator and H2O2 as an enzyme substrate, the DNAzyme catalyzed the reduction of H2O2 to generate quantitative chronoamperometric signal. The catalytic activity of G-quadruplex-Cu2+ DNAzyme for TMB-H2O2 reaction was proportional to the activity of PPase, based on which, a simple and sensitive turn-on electrochemical method for PPase activity was thus developed for the first time. The chronoamperometric intensity of the system had a linear relationship with the PPase activities in the range of 1.0-50.0mU/mL and the detection limit could be down to 0.6mU/mL (S/N = 3). This proposed method was selective, cost-effective and convenient without any labels or complicated operations, which was furthermore applied to screen the inhibitor for PPase with high efficiency.
Copyright © 2017. Published by Elsevier B.V.

KEYWORDS:

Chronoamperometric detection; DNAzyme; G-Quadruplex; Peroxidase-like activity; Pyrophosphatase
PMID:
29136853
DOI:
10.1016/j.talanta.2017.09.069
[Indexed for MEDLINE]
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fredag 6 september 2019

Miten monta PTP fosfataasia osallistuu inositolifosfaattien aineenvaihduntaverkostoon?




 Miksi on niin paljon proteiinityrosiinifosfataaseja,PTP?

 Ihmisen genomissa on  proteiinityrosiinifosfataasigeenejä (PTP)  enemmän kuin proteiinityrosiinikinaasigeenejä (PTK). PTP geenejä on 107 ja  tyrosiinikinaasigeenejä 90. 

PTP-geeneistä on katalyyttisesti inaktiiveja 11 geenia. Kaksi geeniä defosforyloi lähetti-RNA:ta (PIR1 mRNA defosforylaasi; Geeni DUSP11; mRNA capping enzyme, geeni RNGTT)

 ja 13 PTP entsyymiä    toimii inositolifosfolipideihin  (PI)   rajoitetulla alueella  niitä defosforyloimassa ( Koetan löytää tämän joukon:  PTEN geeniryhmään kuuluu 5 geeniä. Myotubulariineista  MTMR ainakin  MTMR2; PTPN9 geenin koodaama PTP-MEG2)

 Täten 81 PTP  entsyymiä  on aktiiveja proteiinifosfataaseja, jotka kykenevät defosforyloimaan  fosfotyrosiinia eli  poistamaan fosfotyrosiinista  (pTyr)  fosfaattia.   

Jos vertaa  proteiinityrosiinikinaasien (PTK)  90 kinaasin suuruiseen  joukkoon , niistä 85 on  katalyyttisesti aktiivia .

Täten  aktiivien tyrosiinifosfataasien (PTP) ja tyrosiinikinaasien (PTK) lukumäärät ovat hyvin samanlaisia. Tästä voisi olettaa että niillä on vertailtavissa olevia substraattispesifisyyksiäkin. 

Kummallakin  entsyymityypillä  on myös  vertailtavissa olevaa  jakaantumistapaa kehossa: yleisestä jakaantumisesta  solutyyppiin rajoittuneeseeen jakaantumiseen siinä määriin, että yksittäiset solut ilmentävät 30%-60%:sti koko PTP-entsyymien kirjoa ja PTK-entsyymien kirjoa. Hermosolut ja vertamuodostavat solut  saattavat ilmentää jopa korkeampia  osuuksia kaikista  proteiinityrosiinifosfataaseista.

Yksittäiset proteiinityrosiinifosfataasit (PTP)  eivät näytä korvaavan toisiansa ja tätä kuvaakin hyvin  raportit  hiirissä esiintyneiden   geenideleetioitten  ainutlaatuisista fenotyypeistä. Esimerkiksi  PTPN22deleetio ( hiirellä PTPN8 deleetio)  aiheuttaa  liiallista T-lymfosyyttimuistisolujen määrän kasvua, pitkittyneitä sekundäärisiä immuunivasteita ja autoimmuniteettia  (Hasegawa et al. 2004). Ihmisellä PTPN22geenin polymorfia korreloi autoimmuunidiabetekseen (Bottini et al. 2004). Muittenkin PTP-geenien mutaatioista seuraa ihmisessä tauteja, mikä  tukee huomiota yksittäisten  proteiinityrosiinifosfataasien  ainutlaatuisista ja tärkeistä toiminnoista..

  Lähde:

Protein Tyrosine Phosphatases in the Human Genome


Open ArchiveDOI:https://doi.org/10.1016/j.cell.2004.05.018

..

 Why So Many PTPs?

The number of genes in the human genome that encode members of the PTP families is higher than anticipated and exceeds the number of genes encoding PTKs (
). However, a direct comparison of the two numbers, 107 and 90, respectively, is not completely fair: of the 107 PTP genes, 11 are catalytically inactive, 2 dephosphorylate mRNA, and 13 dephosphorylate inositol phospholipids. Thus, 81 PTPs are active protein phosphatases with the ability to dephosphorylate phosphotyrosine. Of the 90 PTKs, 85 are through to be catalytically active. Thus, the numbers of active PTPs and PTKs are very similar and one can therefore assume that they have comparable substrate specificities.
 Both types of enzymes also display comparable tissue distribution patterns, from ubiquitous to cell type restricted, so that individual cells express 30%–60% of the entire complement of PTPs and PTKs. Neuronal and hematopoietic cells may express even higher portions of all PTPs.
 That individual PTPs have nonredundant functions is also well illustrated by the unique phenotypes of many of the reported gene deletions in mice. For example, deletion of PTPN22 (PTPN8 in the mouse) causes excessive expansion of memory T lymphocytes, prolonged secondary immune responses, and autoimmunity (). A polymorphism in the human PTPN22 gene also correlates with autoimmune diabetes in humans (). Other human diseases caused by mutations in PTP genes (see discussion below) also support the notion that individual PTPs have unique and important functions.
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ANDREAS ALONSO (2004) Proteiinityrosiinifosfataasiperheitä neljä ja taustageenejä 107 ( Pohdittavanani oleva artikkeli)

https://www.ncbi.nlm.nih.gov/pubmed/15186772


2004 Jun 11;117(6):699-711. Protein tyrosine phosphatases in the human genome.Alonso A1, Sasin J, Bottini N, Friedberg I, Friedberg I, Osterman A, Godzik A, Hunter T, Dixon J, Mustelin T.

Program of Signal Transduction, The Burnham Institute, 10901 North Torrey Pines Road, La Jolla, CA 92037, USA.

Abstract

Tyrosine phosphorylation is catalyzed by protein tyrosine kinases (TKs), which are represented by 90 genes in the human genome.
Here, we present the set of 107 genes in the human genome that encode members of the four protein tyrosine phosphatase (PTP) families.
The four families of PTPases, their substrates, structure, function, regulation, and the role of these enzymes in human disease will be discussed.
PMID:
15186772
DOI:
10.1016/j.cell.2004.05.018

Main Text






 Introduction

Tyrosine phosphorylation is a fundamental mechanism for numerous important aspects of eukaryote physiology, as well as human health and disease ( ,,). Compared to protein phosphorylation in general, phosphorylation on tyrosine is extensively utilized only in multicellular eukaryotes. Tyrosine phosphorylation is used for communication between and within cells, the shape and motility of cells, decisions to proliferate versus differentiate, cellular processes like regulation of gene transcription, mRNA processing, and transport of molecules in or out of cells. Tyrosine phosphorylation also plays an important role in the coordination of these processes among neighboring cells in embryogenesis, organ development, tissue homeostasis, and the immune system. Abnormalities in tyrosine phosphorylation play a role in the pathogenesis of numerous inherited or acquired human diseases from cancer to immune deficiencies.
Although it is generally agreed that tyrosine phosphorylation is regulated by the equal and balanced action of protein tyrosine kinases (PTKs) and protein tyrosine phosphatases (PTPs), proportionately much more research has focused on PTKs. This is partly for historical reasons: the first PTP was purified () and cloned () ten years after the first PTK (). Recent findings have now led to the emerging recognition that PTPs play specific and active, even dominant, roles in setting the levels of tyrosine phosphorylation in cells and in the regulation of many physiological processes (,,,,,).
An important step toward a better appreciation of the PTPs is the clarification of the set of genes in the human genome that encode PTPs. This knowledge will allow for a more global approach to key questions in the PTP field and the regulation of tyrosine phosphorylation-dependent cellular processes in health and disease.

 The 107 Human PTP Genes

Starting with the amino acid sequence of all published PTPs, we used conserved catalytic motifs and whole PTP domains to search the publicly available databases, as well as the SEED database, for genes encoding PTPs or PTP-like genes. Two of our novel PTPs were also found in the Celera and Incyte databases. The genomic locus, exon-intron structure, expression data, and mouse orthologs were clarified for each PTP.
 Previously unpublished PTPs were subjected to a more thorough investigation to verify that they represented bona fide expressed genes with orthologs in the mouse and other organisms.
 Many of these genes have also recently been experimentally verified in our laboratories. The results of this work are a list (Table 1) of all genes in the human genome that encode PTP family members, here defined as all known protein tyrosine phosphatases plus all proteins that contain a domain homologous to the catalytic domain of these known PTPs.
 The list contains 107 genes, 105 of which have a mouse ortholog (one of the missing genes has a rat ortholog). In addition, we estimate that there are at least as many PTP pseudogenes and there are a few bona fide genes (e.g., paladin, KIAA1274) that encode proteins that contain incomplete or distant PTP-like domains. 
Of the 107 human genes, 106 are covered by expressed sequence tags (ESTs). The only gene lacking EST coverage is DUSP27 (DUPD1), which earlier was predicted to encode a phosphatase with an N-terminal cyclophilin-like domain. 
We have been able to amplify a partial cDNA of this gene from testis and muscle mRNA, demonstrating that this gene, too, is expressed. DUSP27 has four exons and is located at 10q22.3 adjacent to DUSP13 at 10q22.2, which in fact consists of two phosphatase genes, which we designate as DUSP13A and DUSP13B (encoding BEDP and TMDP, respectively), which are similar to the catalytic domain of DUSP27. In the mouse genome, the corresponding location on chromosome 14 also contains three PTP genes in tandem, although the sequence of DUSP27 is incomplete. In the chimpanzee genome, there is also a DUSP27 gene, which is 99% identical to the human within the four exons, but differs in the length of some introns. Thus, it appears that an ancestral gene was triplicated (or duplicated twice) before mice and primates diverged during the Cretaceous, 144–165 million years ago.
Table 1The Set of PTP Genes in the Human Genome

Includes all known PTPs in the published literature, verified PTPs or dual-specificity phosphatases in public data bases, and ORFs found by iterative BLAST searches with all the other sequences to find related genes and to assign alternatively spliced forms to the correct genes. To exclude synonyms, splice variants, and incorrectly assembled sequences, a data base with the amino sequences was created. To exclude pseudogenes, EST and other expression data bases were consulted, and the exon-intron structures and chromosomal locations of every gene were determined.
a the human DUSP13 gene contains two DSP genes: DUSP13A (BEDP; 3 exons), two noncoding exons, followed by DUSP13B (TMDP; three exons). In the mouse, the two genes are designated as separate genes.
b formerly DUPD1, but does not contain an N-terminal cyclophilin domain, as originally annotated. The mouse ortholog also lacks cyclophilin domain. Thus, the name DUPD1 is a misnomer.
c we have verified its expression by PCR amplification of a partial cDNA.
d there is a rat homolog, suggesting that a mouse homolog probably does exist.
While the 38 classical PTPs have all been published (reviewed in
), as many as nine of the 107 human PTP genes have not been previously reported (Supplemental Table S1 at http://www.cell.com/cgi/content/full/117/6/699/DC1), although their genomic loci, exon/intron structure, and, in most cases, their homology to PTPs were recognized in databases. These genes include six small phosphatases, DUSP15, DUSP27 (DUPD1), and four genes for which we propose the genomic designations DUSP23, DUSP24, DUSP25, and DUSP26. The protein encoded by DUSP15 is a catalytically active PTP we term VHY (VH1-related member Y;), closely related to VHX encoded by DUSP22. The 150 amino acid residue VHZ (VH1-related member Z) encoded by DUSP25 is the smallest of all catalytically active PTPs (
) and is remarkably well conserved through evolution and even has a 147 amino acid homolog in Thermococcus kodakaraensis, which is 30.3% identical and 49.7% similar over 145 residues. We have verified the existence of a 16 kDa VHZ protein in human cells by immunoblotting. The novel PTPs also include two myotubularin-related proteins, for which we propose the genomic designation MTMR14 and MTMR15. The former is the human ortholog of the Sarcophaga peregrina egg-derived tyrosine phosphatase () and the Drosophila melanogaster gene jumpy (), the mutation of which causes a muscle defect characterized by excessive vibrations of the flight muscles (J.D. and M.J. Wishart, unpublished observation). MTMR15 encodes a catalytically inactive member of the myotubularin group.






 Why So Many PTPs?

The number of genes in the human genome that encode members of the PTP families is higher than anticipated and exceeds the number of genes encoding PTKs (). However, a direct comparison of the two numbers, 107 and 90, respectively, is not completely fair: of the 107 PTP genes,
11 are catalytically inactive, 
2 dephosphorylate mRNA, and
13 dephosphorylate inositol phospholipids. 
Thus, 81 PTPs are active protein phosphatases with the ability to dephosphorylate phosphotyrosine. 
Of the 90 PTKs tyrosine kinases), 85 are through to be catalytically active. Thus, the numbers of active PTPs and PTKs are very similar and one can therefore assume that they have comparable substrate specificities. 
Both types of enzymes also display comparable tissue distribution patterns, from ubiquitous to cell type restricted, so that individual cells express 30%–60% of the entire complement of PTPs and PTKs.
 Neuronal and hematopoietic cells may express even higher portions of all PTPs. That individual PTPs have nonredundant functions is also well illustrated by the unique phenotypes of many of the reported gene deletions in mice. For example, deletion of PTPN22 (PTPN8 in the mouse) causes excessive expansion of memory T lymphocytes, prolonged secondary immune responses, and autoimmunity (). A polymorphism in the human PTPN22 gene also correlates with autoimmune diabetes in humans (). Other human diseases caused by mutations in PTP genes (see discussion below) also support the notion that individual PTPs have unique and important functions.

 Classification of the 107 PTPs into Four Families

Based on the amino acid sequences of their catalytic domains, the PTPs can be grouped into four separate families, each with a range of substrate specificities (Figure 1).
Class I cysteine-based PTPs constitute by far the largest family and contain the 38 well-known “classical” PTPs (), which are strictly tyrosine specific and all have mouse orthologs, and the 61 VH1-like, “dual-specific” protein phosphatases (DSPs), which is the most diverse group in terms of substrate specificity (Figure 1). All Class I enzymes have evolved from a common ancestor, based upon their similar structural folds for classical PTPs, DSPs, and other VH1-like enzymes. 
The single class II cysteine-based PTP in humans is a tyrosine-specific low Mr enzyme, the origin of which appears to be more ancient than class I PTPs: representatives of this family are found in a all major phyla, including plants, numerous prokaryotes, and archea. Class II PTPs are structurally related to bacterial arsenate reductases. 
  Class III cysteine-based PTPs are tyrosine(Y)/threonine(T)-specific phosphatases that most likely evolved from a bacterial rhodanese-like enzyme. They are only represented by the three p80Cdc25 cell cycle regulators. Interestingly, a group of class I PTPs contain a catalytically inactive rhodanese-like domain, referred to as the CDC25 homology (CH2) domain.
 Despite similarities in catalytic mechanism () and active site structure (,,,), class I, II, and III cysteine-based PTPs evolved independently. Nevertheless, a structural comparison indicates that they may have originated from an ancestral fold by rearrangment of elements.
 In contrast, the fourth family of PTPs use a different catalytic mechanism with a key aspartic acid(D)  and dependence on a cation. This family contains the Eya proteins, which were recently discovered to be tyrosine(Y)-, or dual serine (SS)- and tyrosine(Y)-specific protein phosphatases (
,,). Determination of the substrate specificities of the large family of hydrolases to which the Eya PTPs belong will require extensive analysis. Here, only the four Eya proteins are included because they were shown experimentally to be PTPs.






 Class I Cysteine-Based PTPs

The 99 class I cysteine-based family of PTPs can be further classified into several subfamilies based on domain architecture and the degree of homology between catalytic domains. The 38 strictly tyrosine-specific “classical PTPs” can be divided into transmembrane, receptor-like enzymes (RPTPs), and the intracellular, nonreceptor PTPs (NRPTPs). They are represented in the human genome by 21 and 17 genes, respectively (). As is the case for the protein kinases, the number of PTP catalytic domains encoded by the genome is greater than the number of PTP genes because many of the RPTPs have tandem catalytic domains.
 The 61 VH1-like enzymes are much more diverse and can be divided into several subgroups, which share much less sequence identity with each other than the RPTPs do with the NRPTPs. Eleven of the 61 VH1-like PTPs ( MKPs)encoded by the human genome are specific for the mitogen-activated protein (MAP) kinases Erk, Jnk, and p38 (
,,). These MAP kinase phosphatases (MKPs) are characterized by dual phosphothreonine and phosphotyrosine specificity and the presence of a CH2 region and other MAP kinase targeting motifs (,). Another subgroup of DSPs, which we have referred to as the “atypical” DSPs (
), includes a number of poorly characterized enzymes that lack specific MAP kinase targeting motifs and tend to be much smaller enzymes, less than 250 amino acid residues. The first DSP to be cloned, the VH1 protein from Vaccinia virus (), is related to this group, as are the human VHR () and a number of genes given the genomic designations DUSP11, DUS13, DUSP14, DUSP15, DUSP18, DUSP19, DUSP21, DUSP22, and others. 
While VH1 has been reported to dephosphorylate both MAP kinases and Stat1 (), and VHR can dephosphorylate Erk and Jnk in 293T cells () and T cells (,), it appears that many of these small atypical DSP have functions unrelated to MAP kinases. A true outlier is PIR (DUSP11), which dephosphorylates mRNA ().
The three slingshots (SSH1, SSH2, and SSH3) and the three PRLs (PRL-1, PRL-2, and PRL-3) are very poorly understood, while the CDC14 group, which includes KAP, is involved in dephosphorylation of the Cdk activation loop phospho-Thr and inactivation of cyclin-dependent kinases and in exit from mitosis (). 
Finally, the two last subgroups of DSPs, the PTENs (5 genes) and myotubularins (16 genes), have evolved to specifically dephosphorylate the D3-phosphate of inositol phospholipids (). Enzymes with this specificity are present also in yeast. While PTEN dephosphorylates phosphatidylinositol-3,4,5-trisphosphate at the plasma membrane, the myotubularins primarily dephosphorylate phosphatidylinositol-3-phosphate on internal cell membranes (). Both groups also contain catalytically inactive members.

 The Class II Cysteine-Based PTPs

This family is represented in the human genome by a single gene, referred to as ACP1, which encodes the 18 kDa low Mr phosphatase (LMPTP). Related class II enzymes are widely distributed in living organisms with most bacterial genomes encoding at least one member of this family, which are remarkably well conserved through evolution. For example, the human LMPTP is 31% identical to the corresponding protein in Saccharomyces cerevisiae and 39% identical to the YfkJ protein in Bacillus subtilis. Despite the paucity of tyrosine phosphorylation in prokaryotes, the bacterial class II PTPs are bona fide tyrosine-specific phosphatases and in many cases dephosphorylate tyrosine “autokinases” involved in regulation of capsule polysaccharide synthesis. This may represent the ancestral form of tyrosine phosphorylation as a mechanism by which cells sense their extracellular environment, from which receptor PTKs and the counterbalancing PTPs may have evolved. Although the human LMPTP can dephosphorylate a number of tyrosine kinases and their substrates, its physiological function is still unclear. The preservation of a class II PTP through evolution to humans and the correlation of allelic variants of LMPTP with many common human diseases (), such as rheumatoid arthritis, asthma, diabetes, cardiomyopathy, and Alzheimer's disease, indicate that LMPTP likely is involved in the regulation of one or several fundamental processes in cell physiology.

 The Class III Cysteine-Based PTPs

These rhodanese-derived PTPs comprise three cell cycle regulators, CDC25A, CDC25B, and CDC25C, in humans. Their function is to dephosphorylate Cdks at their inhibitory dually phosphorylated N-terminal Thr-Tyr motifs, a reaction that is required for activation of these kinases to drive progression of cells through the cell cycle (
). CDC25s are themselves regulated by phosphorylation. It is curious that a unique type of PTP evolved to serve regulation of the cell cycle, instead of class I or class II enzymes, which presumably already existed at the time. Interestingly, the budding yeast cell cycle can function in the absence of Cdc28 Tyr-15 phosphorylation, and so this layer of regulation may have been a later addition. In other words, CDC25 appears to have entered cell cycle regulation hand in hand with tyrosine phosphorylation of the Cdks. Alternatively, it is possible that an ancestral CDC25 already acted as a rhodanese-type enzyme on Cdks and was transformed into a phosphatase when Cdk tyrosine/threonine phosphorylation evolved. This possibility would better explain why CDC25, rather than an existing PTP, was utilized.

 Asp-Based PTPs

We have listed only the four Eya genes as members of the Asp-based PTPs because they have recently been shown to have Tyr/Ser phosphatase activity (,,). It is clear that this is a much larger family of enzymes, which play important roles in development (,,,), sodium stress in yeast (), and nuclear morphology (). RNA polymerase II C-terminal domain phosphatase is also a member of this family. However, a clear and structurally based definition of this family of enzymes will be needed before the human gene complement of Asp-based phosphatases can be more accurately determined.

 Modular Structure of PTPs

One of the most striking features of the PTP families (Figure 2) is that most enzymes consist of combinations of modular domains. At least 79 of the 107 PTPs contain at least one additional motif or domain (Table 2) outside of their catalytic PTP domain. Many of the domains are protein-protein interaction or phospholipid binding modules. In this respect, PTPs resemble the PTKs (
), but they differ markedly from the serine/threonine-specific protein phosphatases. In PTKs, protein-protein interaction domains serve two distinct purposes: regulation and targeting to substrates and/or subcellular compartments. This appears to be the case also for PTPs, although many of the domains in PTPs are still poorly understood. The set of domains found in PTPs are listed in Table 2 and they include domains that bind specific domains or motifs in other proteins (unphosphorylated or phosphorylated), cellular membranes, the cytoskeleton, or specific phospholipids.
It is interesting to note that the set of protein domains found in PTPs is only partly overlapping with those found in PTKs (). While many PTKs have SH3 and SH2 domains, often in combinations, such as SH3-SH2 or SH2-SH2, only two human PTPs (SHP1 and SHP2) have SH2 domains organized in a tandem fashion like in the Syk family PTKs. Also, the catalytically inactive PTEN-related tensin and C1-TEN have an SH2 domain adjacent to a PTB domain. In contrast, there are no PTPs with SH3 domains. Conversely, there are no known kinases with CRAL/TRIO (Sec14p homology), rhodanese, FYVE, or mRNA capping domains. Nevertheless, it is interesting to note that different domains may serve similar functions in PTPs compared to PTKs. For example, while PTKs mostly use PH domains to interact with phosphoinositides, PTPs use CRAL/TRIO (
) and FYVE domains for the same purpose. Only the myotubularins have PH domains, but it is unclear if they function to interact selectively with phospholipid since myotubularins mostly do not localize to the plasma membrane. In fact, the PH domain of myotubularins was referred to as a GRAM domain until the crystal structure of MTMR2 (
) showed that this region folds as a PH domain.
PTPs also use FERM domains to direct them to the cytoskeleton/plasma membrane interface in a phosphoinositide-dependent manner, analogous to the use of SH3, PH, and C2 domains by kinases. FERM domains may also be able to bind phosphotyrosine, and PTPs may use catalytically inactive PTP (“STYX”) domains in a SH2-like manner to interact with tyrosine phosphorylated proteins (). Together, these differences in domain structure between PTPs and PTKs may reflect the need to regulate these two classes of enzymes in temporally and spatially distinct and often reciprocal manners.

 Hints of Function from the Multidomain Architecture

RPTPs
In many PTPs, the nature of their extracatalytic domains gives some indications as to subcellular location or function. For example, all transmembrane classical PTPs have a membrane-spanning α helix (by definition) and are located in cellular membranes, mostly the plasma membrane, where they interact with the extracellular milieu in a receptor-like fashion. All but RPTPα and RPTPϵ have extended extracellular portions with immunoglobulin, fibronectin, MAM, and carbonic anhydrase domains. Another example is laforin, a PTP that is mutated in an inherited form of progressive myoclonus epilepsy (Lafora's disease;), characterized histologically by accumulation of glycogen-containing granules in the cytoplasm of cells. Recently, laforin was shown to contain a glycogen binding domain, implying a direct role for laforin in glycogen metabolism (
).
 The 17 nonreceptor classical PTPs are particularly rich in protein-protein interaction domains and many of them have several such domains. Within this group, SHP1 and SHP2 provide a good example of how protein-protein interaction domains can cooperate with a PTP domain to achieve both intramolecular regulation and targeting to substrates. In the absence of ligands for the tandem SH2 domain of these PTPs, the more N-terminal SH2 domain folds onto the catalytic domain to block substrate access by the insertion of a loop on the backside of the SH2 domain into the catalytic pocket of the PTP (). When the SH2 domains of this inhibited form of SHP1 or SHP2 encounter a tyrosine phosphorylated ligand, the closed conformation opens and the enzyme is activated some 100-fold (,). Under physiological conditions, SH2 domain ligand binding also juxtaposes the PTP domain to its substrates, which contain, or associate with, the phosphorylated SH2 domain ligand.

The DSP subfamily contains 11 members with a CH2 domain, a region derived from the bacterial rhodanese enzyme and adapted to a MAP kinase docking motif (,
). In CDC25, on the other hand, the rhodanese has evolved into a catalytic PTP domain (). Other noncatalytic domains and motifs found in DSPs include FYVE, glycogen binding, mRNA capping (guanyl methyltransferase), and consensus recognition motifs for N-terminal myristylation or C-terminal prenylation. PTEN contains a C2 domain tightly packed against the PTP domain, while members of the myotubularin family contain PH, FYVE, coiled-coil, and PDZ binding motifs (). A number of additional protein-protein interaction domains are found in PTPs in other organisms, for example a WW domain in a C. elegans PTP (), but are not present in the human PTPs either because they have been lost or because they evolved in a separate lineage.






 Large Multidomain PTPs versus Multisubunit Ser/Thr Phosphatases

The multidomain structure of most PTPs is in sharp contrast to the serine/threonine protein phosphatases, which generally consist of small catalytic subunits that bind regulatory or targeting subunits encoded by separate genes to form a large number of distinct holoenzymes with different biological functions. This dichotomy may explain the differences in gene numbers: 107 PTPs to counteract 90 PTKs versus many fewer catalytic Ser/Thr phosphatase subunits to counter 428 protein kinases and the extensive phosphorylation of more than a third of all cellular proteins. Presumably, the combinatorial subunit principle of serine/threonine protein phosphatases can generate much more diversity and flexibility, but may lack some of the strict specificity and tight regulation possible with single-chain multidomain PTPs. In this context, it may also be significant that the atypical DSP subgroup of PTPs contains many small enzymes devoid of other domains or motifs. It is possible that these enzymes participate in multisubunit complexes where other subunits provide regulation and substrate targeting. However, no examples of this have yet been found.

 Catalytically Inactive PTP Domains

Among the class I cysteine-based PTPs, there are at least 14 catalytically inactive PTP domains, in which critical catalytic residues have been altered. These are the second (D2) domains of CD45, RPTPγ, and RPTPζ, and the only PTP domains of the secretory vesicle-located receptor-like PTP IA-2 (), a VHR-like protein termed STYX (), the MKP-like MK-STYX (), the PTEN-related tensin and C1-ten, and the myotubularin-related proteins MTMR5, MTMR9, MTMR11, MTMR12, MTMR13 (), and MTMR15. Despite lack of catalytic activity, many of these proteins still play important roles in cells, in many cases by partnering with active PTPs. For example, CD45 needs its inactive D2 domain for dephosphorylation by D1 of its physiological substrates (). Similarly, the inactive MTMRs seem to act as important regulatory subunits for active members of the group, as shown by the human disease Charcot-Marie-Tooth type 4B, which is caused by mutation of the catalytically active MTMR2 () or the catalytically inactive MTMR13 (
), in both cases with identical pathology and symptoms. It turns out that these two proteins form a heterodimer, in which the activity of MTMR2 depends on MTMR13. There are similar examples where an inactive kinase domain is needed to activate a catalytically competent kinase domain; e.g., LKB1 is activated by STRAD (). In addition to these inherently inactive PTP domains, alternative splicing of PTP transcripts (which occurs in at least half of all PTPs) can create isoforms that have catalytically inactive PTP domains (
), or lack them altogether (). 
Physiologically important alternative splicing within the catalytic domains is known to create two isoforms of the class II enzyme LMPTP with different specific activity, substrate specificity, and regulation (). In this enzyme, the ratio of the two isoforms is genetically determined and subject to allelic variation, which correlates with the predisposition to several common human diseases ().

 Posttranslational Modifications and Reversible Oxidation of PTPs

The majority of PTPs are posttranslationally modified. Glycosylation appears to be restricted to the transmembrane PTPs, which contain abundant N- and O-linked carbohydrates in their extracellular portions. 
Two groups of DSPs are modified by fatty acids: VHY (DUSP15) and VHX (DUSP22) are N-terminally myristylated in a manner reminiscent of Src family PTKs (
), while the PRLs are C-terminally farnesylated similar to Ras GTPases. By far the most common modification is the phosphorylation of PTPs on serine, threonine, and tyrosine. There are many examples where Ser/Thr phosphorylation has been shown to regulate activity. The phosphorylation of PTPs on tyrosine is particularly intriguing as it implies a physical, if not functional, interaction with PTKs (). At least 15 PTPs have been reported to be tyrosine phosphorylated, but the physiological significance remains unclear in most cases. Tyrosine phosphorylation also introduces the possibility of autodephosphorylation. In fact, many PTPs become noticeably more phosphorylated on tyrosine when expressed as catalytically inactive mutants. In most cases, however, the phosphorylation sites are clearly inaccessible for intramolecular dephosphorylation, suggesting that dephosphorylation must occur in trans. Thus, regulatory networks of PTPs and other protein phosphatases, perhaps in the form of “phosphatase cascades,” may exist in cells.
It has been known for many years that the catalytic cysteine of class I and II PTPs is highly susceptible to oxidation in vitro, and it has been speculated that this could play a role in intact cells exposed to oxidizing agents. An exciting new development was the discovery that oxidation of PTP1B does not result in a stable sulfenic acid derivative of the catalytic site cysteine, but rapidly transforms into a sulfenyl amide ring involving the adjacent serine residue (,
). This form is resistant to further oxidation, which would be irreversible, and is readily reduced back to the free cysteinyl, catalytically active form. Together with recent insights into the production of reactive oxygen species and nitrogen oxides in cells during growth factor stimulation in cancer, inflammation, and neurodegenerative diseases, it now seems likely that reversible redox regulation of PTPs occurs in intact cells. Since PTPs often play dominant roles in setting the levels of tyrosine phosphorylation in cells, this regulation may be physiologically very important.

PTPs as Drug Targets

Compared to the PTKs, many inhibitors (TKI) of which already are in clinical trials, the PTPs are newcomers in the field of drug development. The effect of disruption of the PTP1B gene in mice, which indicated that this PTP acts as a negative regulator of insulin signaling (), ignited the interest of the pharmaceutical industry. Recent discoveries that many other PTPs also play critical roles in a variety of human disease (Table 3) have sparked a growing interest in PTPs as drug targets. In addition to metabolic, neurological, muscle, and autoimmune diseases, at least 30 PTPs have been implicated in cancer (listed in). We expect that with improved understanding of the molecular mechanisms by which PTPs affect the pathophysiology of these diseases, combined with the dominant role that PTPs often play in the regulation of tyrosine phosphorylation-dependent processes, PTP inhibitors will become clinically relevant therapeutics in the future. A rational design of small-molecule inhibitors against PTPs using in silico docking, NMR-based methods, high-throughput crystallization, and specific chemistry will most likely be involved. Following initial concerns about specificity and problems associated with the hydrophilicity of phosphomimetics, promising successes have been achieved by structure-based drug design, particularly those that utilize unique features of the surface topology surrounding the catalytic pocket of each PTP. Figure 3 shows the surface topology and charge distribution of a representative set of catalytic PTP domains to illustrate the striking diversity within the class I PTPs. For example, for PTP1B it was found that a unique second PTyr binding site (,) could be used to develop highly specific bidentate inhibitors that bind both sites (). This general approach can be used to design highly specific and effective inhibitors.

PTPRC (CD45)SCID (Kung et al., 2000;
), multiple sclerosis (
)
PTPRN (IA-2)Antigen for autoimmune diabetes (
PTPRN2 (phogrin)Antigen for autoimmune diabetes
PTPN1 (PTP1B)Insulin resistance, obesity
PTPN6 (SHP1)Sezary syndrome (
PTPN9 (PTP-MEG2)Autism (Smith et al., 2000)
PTPN11 (SHP2)Noonan syndrome
PTPN22 (LYP)SNP polymorphism in type I diabetes
PTEN (PTEN)Bannayan-Zonana (Marsh et al., 1997), Cowden syndrome and Lhermitte-Duclos disease
MTM1 (myotubularin)X-linked myotubular myopathy (
MTMR2 (MTMR2)Charcot-Marie-Tooth syndrome type 4B (
MTMR13 (MTMR13)Charcot-Marie-Tooth syndrome type 4B (
EPM2A (laforin)Progressive myoclonus epilepsy (Lafora's disease) (
ACP1 (LMPTP)Polymorphism correlates with many common diseases (

107 PTP genes 

A. Class I Cys-Based PTPs (99 Genes)
A. 1. Classical PTPs (38 Genes)
A. 1. 1. Transmembrane Classical PTPs (21 Genes)
1. PTPRARPTPα20p13YESYES
2. PTPRBRPTPβ12q15-q21YESYES
3. PTPRCCD45, LCA1q31-q32YESYES
4. PTPRDRPTPδ9p23-p24.3YESYES
5. PTPRERPTPϵ10q26YESYES
6. PTPRFLAR1p34YESYES
7. PTPRGRPTPγ3p21-p14YESYES
8. PTPRHSAP119q13.4YESYES
9. PTPRJDEP1, CD148, RPTPη11p11.2YESYES
10. PTPRKRPTPκ6q22.2-23.1YESYES
11. PTPRMRPTPμ18p11.2YESYES
12. PTPRNIA-2, Islet cell antigen 5122q35-q36.1YESYES
13. PTPRN2PTPRP, RPTPπ, IA-2β, phogrin,7q36YESYES
14. PTPROGLEPP1/PTP-U2/PTPROτ isoforms A/B/C12p13.3-p13.2YESYES
15. PTPRQPTPS3112q21.31YESYES
16. PTPRRPTP-SL, PCPTP,PTPBR7, PC12-PTP112q15YESYES
17. PTPRSRPTPσ19p13.3YESYES
18. PTPRTRPTPρ20q12-q13YESYES
19. PTPRUPTPJ/PTP-U1/PTPRomicron isoforms 1/2/31p35.3-p35.1YESYES
20. PTPRVOST-PTP1q32.1YESYES
21. PTPRZRPTPζ7q31.3YESYES
A. 1.2. NRPTPs (17 Genes),  inracellular


22. PTPN1PTP1B20q13.1-13.2YESYES
23. PTPN2TCPTP, MPTP, PTP-S18p11.3-11.2YESYES
24. PTPN3PTPH19q31YESYES
25. PTPN4PTP-MEG1, TEP2q14.2YESYES
26. PTPN5STEP11p15.1YESYES
27. PTPN6SHP1, PTP1C, SH-PTP1, HCP12p12-13YESYES
28. PTPN7HePTP, LCPTP1q32.1YESYES
29. PTPN9PTP-MEG215q23YESYES
30. PTPN11SHP2, SH-PTP2, Syp, PTP1D, PTP2C, SH-PTP312q24.1YESYES
31. PTPN12PTP-PEST, PTP-P19, PTPG1)7q11.23YESYES
32. PTPN13PTP-BAS, FAP-1, PTP1E, RIP, PTPL1, PTP-BL4q21.3YESYES
33. PTPN14PTP36, PEZ, PTPD21q32.2YESYES
34. PTPN18PTP-HSCF, PTP20, BDP2q21.2YESYES
35. PTPN20TypPTP10q11.22YESYES
36. PTPN21PTPD1, PTP2E, PTP-RL1014q31.3YESYES
37. PTPN22LYP, PEP1p13.3-p13.1YESYES
38. PTPN23HD-PTP, HDPTP, PTP-TD14, KIAA1471, DKFZP564F09233p21.3YESYES
A. 2. DSPs or VH1-like (61 Genes) A. 2. 1. MKPs (11 Genes)


39. DUSP1MKP-1, 3CH134, PTPN10, erp, CL100/ HVH15q34YESYES
40. DUSP2PAC-12q11YESYES
41. DUSP4MKP-2, hVH2/TYP18p12-p11YESYES
42. DUSP5hVH3/B2310q25YESYES
43. DUSP6PYST1, MKP-3/rVH612q22-q23YESYES
44. DUSP7PYST2, B59, MKP-X3p21YESYES
45. DUSP8hVH5, M3/6, HB511p15.5YESYES
46. DUSP9MKP-4, Pyst3Xq28YESYES
47. DUSP10MKP-51q41YESYES
48. DUSP16MKP-7, MKP-M12p13YESYES
49. MK-STYXMK-STYX7q11.23YESYES
A. 2. 2. Atypical DSPs (19 Genes)
50. DUSP3VHR, T-DSP1117q21YESYES
51. DUSP11PIR12p13.1YESYES
52. DUSP12HYVH1, GKAP, LMW-DSP41q21-q22YESYES
53. DUSP13A
BEDP10q22.2YESYES
54. DUSP13B
TMDP, TS-DSP610q22.2YESYES
55. DUSP14MKP6, MKP-L17q12YESYES
56. DUSP15VHY, Q9H1R220q11.21YESYES
57. DUSP18DUSP20, LMW-DSP2022q12.2YESYES
58. DUSP19DUSP17, SKRP1, LDP-2, TS-DSP12q32.1YESYES
59. DUSP21LMW-DSP21, BJ-HCC-26 tumor antigenXp11.4-p11.23YESYES
60. DUSP22VHX, MKPX, JSP1, LMW-DSP2, TS-DSP2, JKAP6p25.3YESYES
61. DUSP23MOSP, similar to RIKEN cDNA 2810004N2011p11.2YESYES
62. DUSP24MGC11368p12YESYES
63. DUSP25VHZ, FLJ20442, LMW-DSP31q23.1YESYES
64. DUSP26VHP, “similar to RIKEN cDNA 0710001B24”2q37.3YESYES
65. DUSP27
DUPD1, FMDSP, “similar to cyclophilin”10q22.3NO
YES
66. EPM2ALaforin6q24YESYES
67. RNGTTmRNA capping enzyme6q16YESYES
68. STYXSTYX14YESYES
A. 2. 3. Slingshots (3 Genes)
69. SSH1SSH1, slingshot 112q24.12YESYES
70. SSH2SSH2, slingshot 217q11.2YESYES
71. SSH3SSH3, slingshot 311q13.1YESYES
A. 2. 4. PRLs (3 Genes)

72. PTP4A1PRL-16q12YESYES
73. PTP4A2PRL-2, OV-11p35YESYES
74. PTP4A3PRL-38q24.3YESYES
A. 2. 5. CDC14s (4 Genes)

75. CDC14ACDC14A1p21YESYES
76. CDC14BCDC14B9q22.33YESYES
77. CDKN3KAP14q22YESYES
78. PTP9Q22PTP9Q229q22.32YESYES
A. 2. 6. PTENs (5 Genes)


79. PTENPTEN, MMAC1, TEP110q23.3YESYES
80. TPIPTPIPα, TPTE and PTEN homologous13q12.11YESYES
81. TPTEPTEN-like, PTEN221p11YESYES
82. TNSTensin2q35-q36YESYES
83. TENC1C1-TEN, TENC1, KIAA107512q13.13YESYES
A. 2. 7. Myotubularins (16 Genes)
84. MTM1myotubularinXq28YESYES
85. MTMR1MTMR1Xq28YESYES
86. MTMR2MTMR211q22YESYES
87. MTMR3MTMR3, FYVE-DSP122q12.2YESYES
88. MTMR4MTMR4, FYVE-DSP217q22-q23YESYES
89. MTMR5MTMR1, SBF122q13.33YESYES
90. MTMR6MTMR613q12YESYES
91. MTMR7MTMR78p22YESYES
92. MTMR8MTMR8Xq11.2YESNO
93. MTMR9MTMR9, LIP-STYX8p23-p22YESYES
94. MTMR10MTMR1015q13.1YESNO
95. MTMR11MTMR11CRA α/β1q12.3YESYES
96. MTMR12MTMR12, 3-PAP5p13.3YESYES
97. MTMR13MTMR13, SBF2, CMT4B211p15.3YESYES
98. MTMR14FLJ22075, hJumpy, hEDTP3p26YESYES
99. MTMR15KIAA101815q13.1YESYES
B. Class II Cys-Based PTPs (1 Gene)
100. ACP1LMPTP, low Mr PTP, LMWPTP, BHPTP2p25YESYES
C. Class III Cys-Based PTPs (3 Genes)
101. CDC25ACDC25A3p21YESYES
102. CDC25BCDC25B20p13YESYES
103. CDC25CCDC25C5q31YESYES
D. Asp-Based PTPs (4 Genes)
104. EYA1Eya18q13.3YESYES
105. EYA1Eya220q13.1YESYES
106. EYA1Eya31p36YESYES
107. EYA1Eya46q23YESYES
107. Total




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