Structural basis for the recognition of a bisphosphorylated MAP kinase peptide by human VHR protein Phosphatase.
Human VHR (vaccinia H1 related phosphatase) is a member of the dual-specificity phosphatases (DSPs) that often act on bisphosphorylated protein substrates. Unlike most DSPs, VHR displays a strong preference for dephosphorylating phosphotyrosine residues over phosphothreonine residues. Here we describe the 2.75 A crystal structure of the C124S inactive VHR mutant in complex with a bisphosphorylated peptide corresponding to the MAP kinase activation lip. This structure and subsequent biochemical studies revealed the basis for the strong preference for hydrolyzing phosphotyrosine within bisphosphorylated substrates containing -pTXpY-. In the structure, the two phospho residues are oriented into distinct pockets; the phosphotyrosine is bound in the exposed yet deep active site cleft while the phosphothreonine is loosely tethered into a nearby basic pocket containing Arg(158). As this structure is the first substrate-enzyme complex reported for the DSP family of enzymes, these results provide the first glimpse into how DSPs bind their protein substrates.
Schumacher
Maria A
MA
Department of Biochemistry and Molecular Biology, Oregon Health and Science University, Portland, Oregon 97201-3098, USA. schumacn@ohsu.edu
Todd
Jacob L
JL
Rice
Adrian E
AE
Tanner
Kirk G
KG
Denu
John M
JM
D001665:Binding Sites; D018360:Crystallography, X-Ray; D054640:Dual Specificity Phosphatase 3; D006801:Humans; D020928:Mitogen-Activated Protein Kinases / Q000737:chemistry* / Q000378:metabolism; D008958:Models, Molecular; D010455:Peptides / Q000737:chemistry; D010766:Phosphorylation; D010769:Phosphothreonine / Q000737:chemistry; D011487:Protein Conformation; D017027:Protein Tyrosine Phosphatases / Q000737:chemistry* / Q000378:metabolism; D013379:Substrate Specificity
D010455:Peptides; D010769:Phosphothreonine; D020928:Mitogen-Activated Protein Kinases; C517025:DUSP3 protein, human; D054640:Dual Specificity Phosphatase 3; D017027:Protein Tyrosine Phosphatases