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IITRS: A Promiscuous Enzyme Synchronizing β-Lactamase with Esterase
Abirlal Mukherjee1, Jyoti Barman1, Rajsekhar Adhikary1
1Department of Biosciences and Bioengineering, Indian Institute of Technology Roorkee, Roorkee 247667, India.
Biochemistry
|October 20, 2025
Summary
A novel penicillin-recognizing protein (IITRS) exhibits dual catalytic activities, acting as both a β-lactamase and esterase. This promiscuous enzyme, found in Enterococcus species, possesses a unique active site distinct from other known enzymes.
Area of Science:
- Biochemistry
- Enzymology
- Microbiology
Background:
- Penicillin-recognizing proteins (PRPs) are crucial in bacterial resistance and drug development.
- Understanding the catalytic mechanisms and substrate specificities of PRPs is essential for designing new therapeutics.
- The discovery of novel PRPs with unique activities can expand our knowledge of enzymatic functions.
Purpose of the Study:
- To identify and characterize a novel penicillin-recognizing protein (IITRS) from Enterococcus species.
- To elucidate the dual catalytic activities (β-lactamase and esterase) of IITRS.
- To investigate the structural and functional basis of IITRS's promiscuity and compare it to known β-lactamases.
Main Methods:
- Isolation and characterization of IITRS from Enterococcus faecium and Enterococcus lactis.
- Site-directed mutagenesis to identify key residues involved in catalysis.
- Enzyme kinetics studies to determine substrate specificity for β-lactams and p-NP esters.
- Inhibition assays using diisopropyl fluorophosphate (DFP).
Main Results:
- IITRS exhibits both β-lactamase and esterase activities, demonstrating catalytic promiscuity.
- The enzyme shows distinct structural features compared to class C β-lactamases, lacking the conserved KTG motif.
- Key residues Ser64 and Tyr150 are involved in β-lactam hydrolysis, with Tyr150 also crucial for esterase activity.
- Activity against p-NP esters increases with longer alkyl chain lengths, and IITRS is inhibited by DFP.
Conclusions:
- IITRS represents a novel class of penicillin-recognizing protein with dual catalytic functions.
- The enzyme possesses a unique active site, distinct from classical β-lactamases and esterases.
- IITRS's promiscuity and unique active site offer potential for biocatalysis, including chiral compound production.
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