The Minimal Essential Core of a Cysteine-based Protein-tyrosine Phosphatase Revealed by a Novel 16-kDa VH1-like Phosphatase, VHZ

Abstract

The smallest active protein-tyrosine phosphatase yet (only 16 kDa) is described here and given the name VHZ for VH1-like member Z because it belongs to the group of small Vaccinia virus VH1-related dual specific phosphatases exemplified by VHR, VHX, and VHY. Human VHZ is remarkably well conserved through evolution as it has species orthologs in frogs, fish, fly, and Archaea. The gene for VHZ, which we designate as DUSP25, is located on human chromosome 1q23.1 and consists of only two coding exons. VHZ is broadly expressed in tissues and cells, including resting blood lymphocytes, Jurkat T cells, HL-60, and RAMOS. In transfected cells, VHZ was located in the cytosol and in other cells also in the nucleoli. Endogenous VHZ showed a similar but more granular distribution. We show that VHZ is an active phosphatase and analyze its structure by computer modeling, which shows that in comparison with the 185-amino acid residue VHR, the 150-residue VHZ is a shortened version of VHR and contains the minimal set of secondary structure elements conserved in all known phosphatases from this class. The surface charge distribution of VHZ differs from that of VHR and is therefore unlikely to dephosphorylate mitogen-activated protein kinases. The remarkably high degree of conservation of VHZ through evolution may indicate a role in some ancient and fundamental physiological process.The VH1-like phosphatases (1) are members of the cysteinebased protein-tyrosine phosphatase (PTP) 1 family and contain the extended consensus signature motif DX 27-30 HCX 2 GX 2 R(S/ T/A)X 5 A(Y/F)LM or slight variations thereof. The crystal structures of the catalytic domains of the VH1-like enzymes VHR (2), mitogen-activated protein kinase phosphatase-3 (MKP-3) (3), KAP (4), PTEN (5), and MTMR2 (6) show that they have the same topology and catalytic machinery as other members of the Class I cysteine-based PTP family exemplified by PTP1B (7) and RPTP␣ (8). However, whereas the catalytic cleft of most VH1-like phosphatases is only 6 Å deep to allow access by the shorter phosphoamino acid side chains of phosphoserine (Ser(P)) and phosphothreonine (Thr(P)) to the catalytic Cys at the bottom of the pocket, the deeper (9 Å) pocket of classical phosphatases like PTP1B only permits dephosphorylation of phosphotyrosine. On the other hand, the catalytic pockets of PTEN (5) and MTMR2 (6) are considerably wider to accommodate their unique physiological substrate, the lipid-bound inositol ring with phosphate at the D position.VH1-like phosphatases are found in all main classes of organisms from bacteria to plants, yeast, insects, worms, and mammals. The human and mouse genomes each encode at least 61 VH1-like enzymes 2 , 11 of which are collectively referred to as the MKPs. In addition to a catalytic VH1-like domain, these MKPs contain a non-catalytic CDC25 homology (CH2) domain for docking with mitogen-activated protein kinases (10), which subsequently are dephosphorylated by the catalytic domain. Another group of 19 small VH1-like phosphatases lack the CH2 doma...

Protein Ser/Thr/Tyr Phosphorylation in the Archaea

“…It appears likely that the handful of archaeal PPMs were acquired "secondhand" from the Bacteria. The widespread phylogenetic dispersal of the cPTPs and LMW PTPs is indicative of ancient origins that likely date back to the last universal common ancestor (33)(34)(35). Although rhodanese, the progenitor of the Cdc25 dual-specificity protein phosphatases, is pervasive among prokaryotic organisms, Cdc25 appears to be exclusively eukaryal in residence and, by extension, origin (24).…”

Dimerization of Vaccinia Virus VH1 Is Essential for Dephosphorylation of STAT1 at Tyrosine 701

“…Thus, the minimal DSP core (aa 28 -171) of VH1 is only minimally active in the absence of its DD. This is in stark contrast to other VH1-like phosphatases like VHZ (also known as DUSP25), for which the minimal ϳ16 kDa DSP core carries all structural and functional determinants sufficient for catalytic activity (33). As a corollary, the minimal sets of secondary structural elements conserved in all knownVH1-like DSPs (5 ␣-helices and 5 ␤-strands) as well as the presence of a catalytic triad (Cys/Arg/ Asp) are not sufficient requirements for a catalytically active DSP core.…”

Identification of Potent VHZ Phosphatase Inhibitors with Structure-Based Virtual Screening

“…Therefore, the inhibitors found in this study seem to be capable of establishing strong multiple hydrogen bonds in the active site of VHZ. Benzoate, benzene-1,2-diol, 5-imino-5,6-dihydro- [1,3,4]thiadiazolo[3,2-a]pyrimidin-7-one, [1,2,4] triazolo [3,4-b] [1,3,4]thiadiazole, nitrobenzene, and 2-hydroxybenzoate moieties in 1, 2 and 5, 3, 4, 6, and 7 are respectively expected to serve as an effective surrogate for the phosphotyrosine group in substrates of VHZ. To the best of our knowledge, compounds 1-7 have not been reported as a phosphatase inhibitor so far, and no additional biological activity was found at least in the two most public chemical databases, ChEMBL and PubChem.…”

“…1 VHZ is one of the smallest phosphatases with 150 amino acids and contains the minimal set of secondary structure elements conserved in all known phosphatases. 2 Recently, it was demonstrated that the activation and the knockdown of VHZ would, respectively, have an effect of enhancing and retarding the progression of the cell cycle associated with carcinogenesis from the G1 to S phase. 3 Furthermore, VHZ was shown to be overexpressed in multiple human primary cancers, including breast cancer.…”