Abstract
We have identified the gene gmk, in the same operon as rpoZ, spoT, and recG at about 82 minutes on the Escherichia coli chromosome. The gmk (GMP kinase) gene encodes a peptide of 23,592 Da, possessing extensive similarity to the amino acid sequence of guanylate kinase from yeast. To confirm that gmk truly encodes guanylate kinase and to explore some of its enzymatic features, we have overproduced the product of gmk and purified it to homogeneity. Unlike guanylate kinases purified from eukaryotic sources, E. coli guanylate kinase is multimeric, and ionic conditions dictate its protomeric state; under low ionic conditions it appears to be a tetramer while under high ionic conditions it is a dimer. Kinetic analysis reveals that guanylate kinase, again, unlike eukaryotic guanylate kinases, binds GMP cooperatively and that the observed cooperatively changes with ionic strength. These results indicate that, despite extensive sequence similarity to its eukaryotic counterparts, E. coli guanylate kinase is structurally and enzymatically different.
Highlights
From the Section on Molecular Regulation, Laboratory of Molecular Genetics, National Institutes of Child Health and Human
Kinetic analysis revealsthat guanylatekinase,again,unlike eukaryotic guanylate kinases, binds GMP cooperatively and that the observed cooperativity changes with ionic strength. These results indicate that, despite extensive sequence similarity to its eukaryotic counterparts, E. coli guanylate kinaseis structurally and enzymatically different
In our study of Guanylate kinases catalyze the reaction (d)GMP ATP the spoT gene of E. coli we have found that gmk
Summary
Kinetic analysis revealsthat guanylatekinase,again,unlike eukaryotic guanylate kinases, binds GMP cooperatively and that the observed cooperativity changes with ionic strength These results indicate that, despite extensive sequence similarity to its eukaryotic counterparts, E. coli guanylate kinaseis structurally and enzymatically different. In our study of Guanylate kinases catalyze the reaction (d)GMP ATP the spoT gene of E. coli we have found that gmk Coli chromosome (9, their importance in maintainingguanine nucleotide pools and 10).The spoT gene encodes the major pyrophosphatase rehaving been purified from a number of sources, only guanylate sponsible for degradingguanosine 3’-5‘ bispyrophosphate kinase from Saccharomyces cerevisiae has been sequenced (2). The crystal structure con- cellular processes by ppGpp or to ppGpp metabolism Firms that it is a monomer with an ATP-binding sitesimilar we report the sequence of E. coli gmk as well as the overproto thatof adenylate kinase and other ATP- and GTP-bindingduction, purification, and biochemical characterization of its proteins. E. coli guanylate kinasewere aligned with the FASTP program (23)
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