Structural insights into the prereaction state of pyruvate decarboxylase from Zymomonas mobilis .
Pyruvate decarboxylase (PDC) uses thiamine diphosphate as an essential cofactor to catalyze the formation of acetaldehyde on the pathway of ethanol synthesis. Here we report the crystallographic image of a prereaction intermediate of a bacterial pyruvate decarboxylase prepared by cocrystallizing the enzyme with pyruvate and a stable analogue of the cofactor's activated ylid form. A second crystal structure of PDC in complex with fluoride shows that the ion organizes a water molecule that occludes the pyruvate binding site, accounting for the inhibitory effect of the halide. Also reported is a structure of the cofactor-free apo form, which when compared to the structure of the holo form indicates how thiamine diphosphate organizes the active site pocket of pyruvate decarboxylase to support catalysis. Guided by the structural and enzymatic data, we propose roles for several key residues in the catalytic mechanism.
Pei
Xue-Yuan
XY
Department of Biochemistry, University of Cambridge, Tennis Court Road, Cambridge CB2 1QA, UK.
Erixon
Karl M
KM
Luisi
Ben F
BF
Leeper
Finian J
FJ
D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D005459:Fluorides / Q000737:chemistry / Q000378:metabolism; D017433:Protein Structure, Secondary; D011767:Pyruvate Decarboxylase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D019289:Pyruvic Acid / Q000737:chemistry / Q000378:metabolism; D017355:Zymomonas / Q000201:enzymology*
D001426:Bacterial Proteins; D019289:Pyruvic Acid; D011767:Pyruvate Decarboxylase; D005459:Fluorides