The partial amino acid sequence of yeast 3-phosphogly cerate kinase
Yeast 3-phosphoglycerate kinase (PGK), a glycolytic enzyme, was isolated, purified and characterised, and subjected to cyanogen bromide cleavage. Three of the four cyanogen bromide fragments were purified by gel filtration, and enzymically digested with trypsin, chymotrypsin, pepsin, thermolysin and staph. protease. The resulting peptides were purified by high voltage electrophoresis on paper, and were sequenced by the dansyl-Edman technique. Of the 243 amino acids constituting these three cyanogen bromide fragments, 221 were placed in order by direct sequencing, composition, and comparison with the horse muscle enzyme sequence, with which yeast PGK exhibits an estimated 65% identity. By fitting the sequence to the 2.5 R-resolution electron density map of yeast PGK, the positions of the reactive alutamyl residue (Glu-403) and of the reactive tyrosyl residue (Tyr-195) have been located, at sites distant from that of bound ATP. The remote location of the glutamyl residue supports the suggestion that PGK must undergo a substrate-induced conformational change in order to bring Glu-403 into closer proximity with the ~-phosphoryl of ATP. The glycine lining of the ATP-binding pocket of the horse muscle enzyme is conserved in the yeast enzyme, as are the glycines near the t-phosphoryl of bound ATP. However, the location of the hinge region of the molecule, tentatively identified in the horse muscle enzyme sequence, is not conserved in the yeast PGK sequence. The structure of PGK has been compared with that of other kinases, and similarities with hexokinase and adenylate kinase noted.