Mapping catalytically important regions of an enzyme using two-codon insertion mutagenesis: a case study correlating beta-lactamase mutants with the three-dimensional structure.

Overview

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.