Characterizing Two Cytochrome P450s in Tiacumicin Biosynthesis Reveals Reaction Timing for Tailoring Modifications

Abstract

Functions of the tailoring enzymes for the biosynthesis of tiacumicin B (1) have been previously determined. However, the reaction timing remains elusive. Herein, we report the in vitro biochemical characterization of two P450 enzymes TiaP1 and TiaP2. The investigation of their substrate scope and kinetic parameters clearly demonstrates that the TiaP2-catalyzed C-20 hydroxylation is the first tailoring step and the TiaP1-catalyzed C-18 hydroxylation is the last step in the biosynthesis of 1.

Chemistry and Biology of the Clinically Used Macrolactone Antibiotic Fidaxomicin

“…hamdenensis , one of the known producer strains . According to sequence analysis and analysis of truncated products isolated from the fermentation broth of gene‐knockout mutants a possible biosynthetic pathway was proposed …”

“…The complete catalytic order of the assembly of the final natural product 1 remains elusive. However, latest investigations indicated that TiaP2 first oxidizes macrocycle 32 at the C(20)‐position, before glycosyltransferase TiaG1 attaches the noviose to form alcohol 42 . Then, the isobutyric ester is attached by acyltransferase TiaS6 or rhamnosylation occurs to construct fidaxomicin scaffold 43 .…”

Novel Fidaxomicin Antibiotics through Site-Selective Catalysis

“…18,22,23 So far, most known derivatives were obtained through fermentation of gene-knockout mutants of the producer strain. [24][25][26][27] Furthermore, several compounds with a similar scaffold were found along with fidaxomicin (1), some of which lack the rhamnoseorsellinate moiety and/or the noviose. 8,28 Those truncated products missing one of the sugar moieties display significantly decreased antibiotic activity and thereby point out the importance of these carbohydrate units.…”

Novel fidaxomicin antibiotics through site-selective catalysis

“…Although fidaxomicin has a high potential to deliver next-generation antibiotics, there are only few successful approaches toward new derivatives documented that might broaden structure-activity relationship (SAR) information and thereby further extend its application 18,28,29 . So far, most known derivatives were obtained through fermentation of geneknockout mutants of the producer strain or by chemical modification of the homoorsellinic acid moiety [30][31][32][33][34][35] . Furthermore, several compounds with a similar scaffold were found along with fidaxomicin (1), some of which lack the rhamnose-orsellinate and/or the noviose moiety 8,36 .…”