The importance of arginine 102 for the substrate specificity of Escherichia coli malate dehydrogenase.
The malate dehydrogenase from Escherichia coli has been specifically altered at a single amino acid residue by using site-directed mutagenesis. The conserved Arg residue at amino acid position 102 in the putative substrate binding site was replaced with a Gln residue. The result was the loss of the high degree of specificity for oxaloacetate. The difference in relative binding energy for oxaloacetate amounted to about 7 kcal/mol and a difference in specificity between oxaloacetate and pyruvate of 8 orders of magnitude between the wild-type and mutant enzymes. These differences may be explained by the large hydration potential of Arg and the formation of a salt bridge with a carboxylate group of oxaloacetate.
Nicholls
D J
DJ
Division of Biotechnology, PHLS Centre for Applied Microbiology and Research, Porton Down, Salisbury, UK.
Miller
J
J
Scawen
M D
MD
Clarke
A R
AR
Holbrook
J J
JJ
Atkinson
T
T
Goward
C R
CR
D000595:Amino Acid Sequence; D001120:Arginine; D001665:Binding Sites; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D005973:Glutamine; D007700:Kinetics; D008291:Malate Dehydrogenase / Q000235:genetics / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D013379:Substrate Specificity
D005973:Glutamine; D001120:Arginine; D008291:Malate Dehydrogenase