Premium
Activator anion binding site in pyridoxal phosphorylase b : The binding of phosphite, phosphate, and fluorophosphate in the crystal
Author(s) -
Oikonomakos Nikos G.,
Zographos Spyros E.,
Tsitsanou Katerina E.,
Johnson Louise N.,
Acharya K. Ravi
Publication year - 1996
Publication title -
protein science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.353
H-Index - 175
eISSN - 1469-896X
pISSN - 0961-8368
DOI - 10.1002/pro.5560051204
Subject(s) - glycogen phosphorylase , chemistry , activator (genetics) , pyridoxal phosphate , phosphate , binding site , pyridoxal 5 phosphate , pyridoxal , biochemistry , stereochemistry , enzyme , cofactor , receptor
It has been established that phosphate analogues can activate glycogen phosphorylase reconstituted with pyridoxal in place of the natural cofactor pyridoxal 5′‐phosphate (Chang YC, McCalmont T, Graves DJ. 1983. Biochemistry 22 :4987–4993). Pyridoxal phosphorylase b has been studied by kinetic, ultracentrifugation, and X‐ray crystallographic experiments. In solution, the catalytically active species of pyridoxal phosphorylase b adopts a conformation that is more R‐state‐like than that of native phosphorylase b , but an inactive dimeric species of the enzyme can be stabilized by activator phosphite in combination with the T‐state inhibitor glucose. Co‐crystals of pyridoxal phosphorylase b complexed with either phosphite, phosphate, or fluorophosphate, the inhibitor glucose, and the weak activator IMP were grown in space group P 4 3 2 1 2, with native‐like unit cell dimensions, and the structures of the complexes have been refined to give crystallographic R factors of 18.5–19.2%, for data between 8 and 2.4 Å resolution. The anions bind tightly at the catalytic site in a similar but not identical position to that occupied by the cofactor 5′‐phosphate group in the native enzyme (phosphorus to phosphorus atoms distance = 1.2 Å). The structural results show that the structures of the pyridoxal phosphorylase b ‐anion‐glucose‐IMP complexes are overall similar to the glucose complex of native T‐state phosphorylase b . Structural comparisons suggest that the bound anions, in the position observed in the crystal, might have a structural role for effective catalysis.