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Molecular characterization of three chloramphenicol acetyltransferases isolated from Haemophilus influenzae
Insights
Three chloramphenicol acetyltransferase enzymes from Haemophilus influenzae were characterized. These enzymes are related to the type II group but show distinct molecular and activity differences.
Area of Science:
- Microbiology
- Enzymology
- Molecular Biology
Background:
- Plasmid-mediated antibiotic resistance is a significant concern in bacterial infections.
- Chloramphenicol acetyltransferases (CATs) are key enzymes conferring resistance to chloramphenicol.
- Haemophilus influenzae is an important human pathogen where resistance mechanisms are actively studied.
Purpose of the Study:
- To purify and characterize three plasmid-mediated chloramphenicol acetyltransferases (CATs) from Haemophilus influenzae.
- To compare the properties of these enzymes with known CAT types, particularly the enteric type II enzyme.
- To elucidate the relationship between CAT enzymes from different Haemophilus influenzae strains.
Main Methods:
- Purification of three distinct plasmid-mediated chloramphenicol acetyltransferases.
- Biochemical characterization including sensitivity to 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB).
- Chromatographic analysis using a chloramphenicol base-bound resin and immunological comparison via antisera.
Main Results:
- All three Haemophilus influenzae CAT enzymes exhibited properties consistent with the gram-negative family of CATs.
- Enzymes showed sensitivity to DTNB, similar elution profiles, and comparable reactions to antisera as the enteric type II enzyme.
- Differences were observed in subunit molecular weight, enzyme activity, and partial protein digestion patterns among the four enzymes.
Conclusions:
- The three characterized Haemophilus influenzae CAT enzymes belong to the less common type II group.
- These enzymes are related to each other and to the enteric type II enzyme, but are not identical.
- The findings contribute to understanding the diversity and evolution of antibiotic resistance mechanisms in Haemophilus influenzae.
Abstract:
Three plasmid-mediated chloramphenicol acetyltransferases isolated from different Haemophilus influenzae strains were purified and characterized. All three enzymes had properties in common with the gram-negative family of chloramphenicol acetyltransferase. The Haemophilus enzymes and the enteric type II enzyme were sensitive to 5,5'-dithiobis(2-nitrobenzoic acid), gave the same elution patterns from a highly substituted resin containing a bound chloramphenicol base, and had similar reactions to antisera. All four differed from each other in subunit molecular weight, enzyme activity, and partial protein digestion patterns. The data suggest that the three Haemophilus enzymes belong to the less common type II group and are related, but is not identical, to each other and to the enteric type II enzyme.