The high resolution crystal structure of yeast hexokinase PII with the correct primary sequence provides new insights into its mechanism of action

Hexokinase is the first enzyme in the glycolytic pathway, catalyzing the tr ansfer of a phosphoryl group from ATP to glucose to form glucose 6-phosphat e and ADP. Two yeast hexokinase isozymes are known, namely PI and PII. The crystal structure of yeast hexokinase PII from Saccharomyces cerevisiae wit hout substrate or competitive inhibitor is determined and refined in a tetr agonal crystal form at 2.2-Angstrom resolution The folding of the peptide c hain is very similar to that of Schistosoma mansoni and previous yeast hexo kinase models despite only 30% sequence identity between them. Distinct dif ferences in conformation are found that account for the absence of glucose in the binding site. Comparison of the current model with S. mansoni and ye ast hexokinase PI structures both complexed with glucose shows in atomic de tail the rigid body domain closure and specific loop movements as glucose b inds. A hydrophobic channel formed by strictly conserved hydrophobic residu es in the small domain of the hexokinase is identified. The channel's mouth is close to the active site and passes through the small domain to its sur face. The possible role of the observed channel in proton transfer is discu ssed.