Open AccessGuanylate kinase from Saccharomyces cerevisiae
Author(s) -
BERGER Albin,
SCHILTZ Emile,
SCHULZ Georg E.
Publication year - 1989
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1989.tb15035.x
Subject(s) - adenylate kinase , biochemistry , cysteine , saccharomyces cerevisiae , kinase , size exclusion chromatography , enzyme , chemistry , peptide sequence , amino acid , yeast , gene
This paper describes a large‐scale purification of guanylate kinase (ATP + GMP · ADP + GDP) from Saccharomyces cerevisiae , the crystallization of the enzyme and preliminary X‐ray investigations. Furthermore the complete amino acid sequence of the enzyme has been determined and was compared to adenylate kinase sequences.1 Guanylate kinase was purified in five steps to homogeneity: crude extract, ion‐exchange chromatography, affinity chromatography and gel filtration twice. 2 The enzyme was crystallized to single octahedral bipyramids with sizes up to 500 × 200 × 150 μm 3 . Preliminary X‐ray results are given. 3 The final sequence shows 186 amino acids ( M r = 20 548), containing one cysteine and one tryptophan. It was determined from peptides of five cleavages of the whole protein. Three cleavages were used for determination of the whole polypeptide chain. From the other two, only some peptides were used to secure overlaps and the cysteine position. The N‐terminal blocking group was identified by 1 H‐NMR spectroscopy. 4 Since guanylate kinase shows the mononucleotide binding pattern GXXGXGK, it was compared to other proteins containing this pattern. But no further homology signal could be detected. A comparison with adenylate kinases revealed significant similarity in another chain segment. This led to the conclusion that guanylate kinase is at least partially homologous to the adenylate kinases.