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Substrate specificity of isopenicillin N synthase.
G W Huffman1, P D Gesellchen, J R Turner
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana 46285.
Journal of Medicinal Chemistry
|May 15, 1992
Summary
Synthetic modifications of the isopenicillin N synthase (IPNS) substrate (ACV) were tested. While some analogues induced beta-lactamase, none matched ACV's efficiency, and certain modifications yielded inhibitors.
Area of Science:
- Biochemistry
- Enzymology
- Medicinal Chemistry
Background:
- Isopenicillin N synthase (IPNS) is crucial for penicillin biosynthesis.
- The natural substrate for IPNS is (L-alpha-amino-delta-adipyl)-L-cysteinyl-D-valine (ACV).
- Understanding substrate specificity can guide the development of novel antibiotics.
Purpose of the Study:
- To investigate the substrate specificity of IPNS using synthetic ACV analogues.
- To determine the impact of specific amino acid modifications on IPNS activity.
- To evaluate the potential of modified tripeptides as substrates or inhibitors.
Main Methods:
- Purification of IPNS from Penicillium chrysogenum and recombinant Escherichia coli.
- In vitro assays using synthetic ACV analogues and beta-lactamase induction.
- Synthesis and antibacterial evaluation of a beta-lactam product from an ACV analogue.
Main Results:
- Various amino and carboxyl terminal modifications of ACV elicited beta-lactamase induction.
- No ACV analogue demonstrated substrate efficiency comparable to the natural ACV.
- Modifications at the L-cysteine or D-valine positions abolished substrate activity or created inhibitors.
Conclusions:
- The natural substrate ACV is highly optimized for IPNS activity.
- Specific modifications, particularly at the D-valine residue, can transform analogues into potent IPNS inhibitors.
- These findings offer insights into enzyme-substrate interactions and potential drug design strategies.