Crystal structure of the yeast His6 enzyme suggests a reaction mechanism.
The Saccharomyces cerevisiae His6 gene codes for the enzyme phosphoribosyl-5-amino-1-phosphoribosyl-4-imidazolecarboxamide isomerase, catalyzing the fourth step in histidine biosynthesis. To get an insight into the structure and function of this enzyme, we determined its X-ray structure at a resolution of 1.30 A using the anomalous diffraction signal of the protein's sulphur atoms at 1.77 A wavelength. His6 folds in an (alpha/beta)8 barrel similar to HisA, which performs the same function in bacteria and archaea. We found a citrate molecule from the buffer bound in a pocket near the expected position of the active site and used it to model the open form of the substrate (phosphoribulosyl moiety), which is a reaction intermediate. This model enables us to identify catalytic residues and to propose a reaction mechanism where two aspartates act as acid/base catalysts: Asp134 as a proton donor for ring opening, and Asp9 as a proton acceptor and donor during enolization of the aminoaldose. Asp9 is conserved in yeast His6 and bacterial or archaeal HisA sequences, and Asp134 has equivalents in both HisA and TrpF, but they occur at a different position in the protein sequence.
Quevillon-Cheruel
Sophie
S
Institut de Biochimie et de Biophysique Moléculaire et Cellulaire (CNRS-UMR 8619), Université Paris-Sud, 91405 Orsay, France.
Leulliot
Nicolas
N
Graille
Marc
M
Blondeau
Karine
K
Janin
Joel
J
van Tilbeurgh
Herman
H
D019747:Aldose-Ketose Isomerases / Q000737:chemistry* / Q000378:metabolism*; D000619:Aminohydrolases / Q000737:chemistry / Q000378:metabolism; D001224:Aspartic Acid / Q000378:metabolism; D001665:Binding Sites; D020134:Catalytic Domain; D002951:Citrates / Q000378:metabolism; D018360:Crystallography, X-Ray / Q000379:methods; D008958:Models, Molecular; D011487:Protein Conformation; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000378:metabolism; D013455:Sulfur / Q000737:chemistry
D002951:Citrates; D029701:Saccharomyces cerevisiae Proteins; D001224:Aspartic Acid; D013455:Sulfur; C087227:imidazole glycerol phosphate synthase; D000619:Aminohydrolases; D019747:Aldose-Ketose Isomerases; C079656:Phosphoribosyl-5-amino-1-phosphoribosyl-4-imidazolecarboxiamide isomerase