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Experimental and Computational Studies on the Biotransformation of Pseudopyronines with Human Cytochrome P450 CYP4F2
Ya Lu1, Xueling Liu1,2, Rowaa Lotfy3
1School of Pharmaceutical Science and Technology, Tianjin University, 92 Weijin Road, Tianjin 300092, People's Republic of China.
Abstract:
The secondary metabolite pseudopyronine B, isolated from Pseudomonas mosselii P33, was biotransformed by human P450 enzymes, heterologously expressed in the fission yeast Schizosaccharomyces pombe. Small-scale studies confirmed that both CYP4F2 and CYP4F3A were capable of oxidizing the substrate, with the former achieving a higher yield. In larger-scale studies using CYP4F2, three new oxidation products were obtained, the structures of which were elucidated by UV-vis, 1D and 2D NMR, and HR-MS spectroscopy. These corresponded to hydroxylated, carboxylated, and ester derivatives (1-3) of pseudopyronine B, all of which had been oxidized exclusively at the ω-position of the C-6 alkyl chain. In silico homology modeling experiments highlighted key interactions between oxygen atoms of the pyrone ring and two serine residues and a histidine residue of CYP4F2, which hold the substrate in a suitable orientation for oxidation at the terminus of the C-6 alkyl chain. Additional modeling studies with all three pseudopyronines revealed that the seven-carbon alkyl chain of pseudopyronine B was the perfect length for oxidation, with the terminal carbon lying close to the heme iron. The antibacterial activity of the substrates and three oxidation products was also assessed, revealing that oxidation at the ω-position removes all antimicrobial activity. This study both increases the range of known substrates for human CYF4F2 and CYP4F3A enzymes and demonstrates their utility in producing additional natural product derivatives.
Insights
Human P450 enzymes, specifically CYP4F2, biotransformed pseudopyronine B into new derivatives. This oxidation at the alkyl chain
Area of Science:
- Biochemistry
- Natural Product Chemistry
- Enzymology
Background:
- Pseudopyronine B is a secondary metabolite from *Pseudomonas mosselii*.
- Human P450 enzymes are crucial for metabolizing various compounds.
- Heterologous expression systems are valuable for studying enzyme function.
Purpose of the Study:
- To investigate the biotransformation of pseudopyronine B by human P450 enzymes.
- To identify and characterize the oxidation products of pseudopyronine B.
- To explore the potential of P450 enzymes in natural product derivative synthesis.
Main Methods:
- Biotransformation of pseudopyronine B using heterologously expressed CYP4F2 and CYP4F3A in *Schizosaccharomyces pombe*.
- Structure elucidation of oxidation products using UV-vis, NMR, and HR-MS.
- In silico homology modeling to understand enzyme-substrate interactions.
- Assessment of antibacterial activity of substrates and products.
Main Results:
- CYP4F2 and CYP4F3A oxidized pseudopyronine B, with CYP4F2 yielding higher product amounts.
- Three new derivatives (hydroxylated, carboxylated, esterified) were identified, oxidized at the ω-position of the C-6 alkyl chain.
- Homology modeling revealed specific interactions positioning pseudopyronine B for terminal oxidation.
- Oxidation at the ω-position abolished the antibacterial activity of pseudopyronine B.
Conclusions:
- Human CYP4F2 and CYP4F3A enzymes can biotransform pseudopyronine B.
- The study expands the known substrate range for these P450 enzymes.
- Enzymatic oxidation provides a route to novel pseudopyronine B derivatives with altered biological activity.
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