Mapping catalytically important regions of an enzyme using two-codon insertion mutagenesis: a case study correlating beta-lactamase mutants with the three-dimensional structure.
Two-codon insertion mutants throughout the beta-lactamase (Bla)-encoding gene were characterized. Second site revertants of various mutants were isolated, mapped and sequenced. The activity of the mutants and the ability to get revertants showed a positive correlation with increasing distance from the active site, based on the three-dimensional structure of Bla. This observation is discussed as it may pertain to the generalized use of two-codon insertion mutagenesis in mapping important catalytic regions in enzymes.
Zebala
J
J
Department of Microbiology, Hearst Microbiology Research Center, Cornell University Medical College, New York, NY 10021.
Barany
F
F
D000595:Amino Acid Sequence; D001407:Bacillus / Q000201:enzymology; D003062:Codon / Q000235:genetics; D005798:Genes, Bacterial; D008958:Models, Molecular; D016254:Mutagenesis, Insertional; D010957:Plasmids; D011487:Protein Conformation; D015183:Restriction Mapping; D013211:Staphylococcus aureus / Q000201:enzymology; D013302:Streptomyces / Q000201:enzymology; D001618:beta-Lactamases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
D003062:Codon; D001618:beta-Lactamases