Intramolecular interactions in protein tyrosine phosphatase RPTPmu: kinetic evidence.
The receptor-like protein tyrosine phosphatase RPTPmu contains three intracellular domains: the juxtamembrane (JM) and two phosphatase domains (D1 and D2). D1 is catalytically active in vitro. The functional roles of JM and D2 are still unclear. To find out whether and how they modulate the phosphatase activity of D1, we compared the enzymatic characteristics of two constructs, containing a truncated JM and either D1 or both phosphatase domains. p-Nitrophenyl phosphate and two peptide substrates were efficiently dephosphorylated by both constructs. The specificity constant of D1 alone was up to 50% higher. D2 induces (a) decreased K(m) values for peptide substrates, (b) decreased catalytic efficiency for these substrates, (c) shifting of the optimal pH to slightly lower values, and (d) looser binding of competitive inhibitors. These data suggest that the phosphatase activity of D1 is negatively modulated and its ligand binding capacity is sensibly modified by domain D2, having possible functional significance.
Aricescu
A R
AR
Department of Enzymology, Institute of Biochemistry, Splaiul Independentei 296, 77700 Bucharest, Romania.
Fulga
T A
TA
Cismasiu
V
V
Goody
R S
RS
Szedlacsek
S E
SE
D000595:Amino Acid Sequence; D002384:Catalysis; D020134:Catalytic Domain; D002462:Cell Membrane / Q000201:enzymology; D003001:Cloning, Molecular; D004795:Enzyme Stability; D005755:Gastrins / Q000737:chemistry; D006629:Hirudins / Q000737:chemistry; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D010748:Phosphopeptides / Q000737:chemistry* / Q000378:metabolism; D010744:Phosphoric Monoester Hydrolases / Q000737:chemistry / Q000378:metabolism; D019000:Phosphotyrosine; D017027:Protein Tyrosine Phosphatases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D054623:Receptor-Like Protein Tyrosine Phosphatases, Class 2; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D015183:Restriction Mapping; D013379:Substrate Specificity
D005755:Gastrins; D006629:Hirudins; D010446:Peptide Fragments; D010748:Phosphopeptides; D011993:Recombinant Fusion Proteins; D019000:Phosphotyrosine; D010744:Phosphoric Monoester Hydrolases; C516859:PTPRM protein, human; D017027:Protein Tyrosine Phosphatases; D054623:Receptor-Like Protein Tyrosine Phosphatases, Class 2