Crystal structures of a low-molecular weight protein tyrosine phosphatase from Saccharomyces cerevisiae and its complex with the substrate p-nitrophenyl phosphate

Low-molecular weight protein tyrosine phosphatases are virtually ubiquitous , which implies that they have important cellular functions. We present her e the 2.2 Angstrom resolution X-ray crystallographic structure of wild-type LTP1, a low-molecular weight protein tyrosine phosphatase from Saccharomyc es cerevisiae. We also present the structure of an inactive mutant substrat e complex of LTP1 with p-nitrophenyl phosphate (pNPP) at a resolution of 1. 7 Angstrom. The crystal structures of the wild-type protein and of the inac tive mutant both have two molecules per asymmetric unit. The wild-type prot ein crystal was grown in HEPES buffer, a sulfonate anion that resembles the phosphate substrate, and a HEPES molecule was found with nearly full occup ancy in the active site. Although the fold of LTP1 resembles that of its bo vine counterpart BPTP, there are significant changes around the active site that explain differences in their kinetic behavior. In the crystal of the inactive mutant of LTP1, one molecule has a pNPP in the active site, while the other has a phosphate ion. The aromatic residues lining the walls of th e active site cavity exhibit large relative movements between the two molec ules. The phosphate groups present in the structures of the mutant protein bind more deeply in the active site (that is, closer to the position of nuc leophilic cysteine side chain) than does the sulfonate group of the HEPES m olecule in the wild-type structure. This further confirms the important rol e of the phosphate-binding loop in stabilizing the deep binding position of the phosphate group, thus helping to bring the phosphate close to the thio late anion of nucleophilic cysteine, and facilitating the formation of the phosphoenzyme intermediate.