National Center for
4L8W: Crystal Structure Of Gamma Glutamyl Hydrolase (h218n) From Zebrafish Complex With Mtx Polyglutamate
Structural insights into the hydrolysis and polymorphism of methotrexate polyglutamate by zebrafish gamma-glutamyl hydrolase
J. Med. Chem. (2013) 56 p.7625-7635
gamma-Glutamyl hydrolases (gammaGH) catalyze the hydrolysis of gamma-linked glutamate residues from the polyglutamyl of folates and antifolates, such as methotrexate (MTX), a widely used anticancer drug. We describe the first crystal structures of the endopeptidase-type gammaGH (zgammaGH) from zebrafish and the mutant complexes with MTX(Glu)5 and hydrolyzed MTX(Glu)1, revealing the complete set of key residues involved in hydrolysis as well as the substrate-binding subsites (-1 to +2). The side chain of Phe20 and the 6-methylpterin ring of MTX(Glu)5 invoke pi-pi interactions to promote distinct concerted conformational alterations involving approximately 90 degrees rotations in the complexes with the zgammaGH-C108A and zgammaGH-H218N mutant proteins. The structural geometries of the MTX(Glu)5 and hydrolyzed MTX(Glu)1 in the mutant complexes differ significantly from those of the previously known MTX(Glu)1, providing polymorphic information. Together with the structural comparison and the activity analysis, these results shed light on the catalytic mechanism and substrate recognition of zgammaGH and other gamma-glutamyl hydrolases.