Investigations of carbocyclisation reactions in prodiginine alkaloid biosynthesis

• Main Author: Salih, Rebin M.
• Published: University of Warwick 2017
• Subjects: 540; QD Chemistry
• Online Access: https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.752484

Prodiginine alkaloids, including streptorubin B, metacycloprodigiosin and roseophilin, are a family of red-pigmented pyrrole-containing specialised metabolites with immunosuppressive, anticancer and antimalarial properties. The final steps in streptorubin B and metacycloprodigiosn biosynthesis are oxidative carbocyclisations catalysed by Rieske oxygenases RedG and McpG respectively. These enzymes utilize the same substrate (undecylprodigiosin) to give cyclised products with different regio and stereochemistry. In order to further investigate the McpG catalysed reaction and to develop a system that could be used to probe the mechanism, mcpG was expressed in S. albus and S. coelicolor M511. Cyclic compound was observed following feeding of undecylprodigiosin to S. albus/mcpG. Since production was low, the role of McpH in forming a complex with McpG was investigated. Unfortunately, whilst McpH was shown to catalyse the condensation of 2-undeyclpyrrole (2-UP) and 4-methoxy-2, 2’ bipyrrole-5-carboxaldehyde (MBC), co-expression of mcpH with mcpG did not improve production of metacycloprodigiosin. Since whole genome sequencing showed that the mcpG expression construct originally used to prove the function of this gene contained a 5' truncation due to a sequencing error, full length mcpG was expressed in a redG mutant of S. coelicolor. This resulted in production of metacyloprodigiosin, which was confirmed by mass spectrometry and 1H and 13C NMR spectroscopy. The rph gene cluster reported to be responsible for the biosynthesis of prodigiosin R1 and roseophilin in S. griseoviridis contains four redG orthologues (rphG, rphG2, rphG3 and rphG4). Unfortunately, neither expression of the rphG genes in the redG mutant of S. coelicolor, nor the feeding of 11-methyldodecylprodigiosin to S. albus separately expressing each of the rphG genes resulted in the production of carbocylic products. Bioinformatics analysis of the S. longispororuber genome sequence revealed several cryptic natural product biosynthetic gene clusters. The likely products of some of these clusters are discussed in the light of predictive sequence analysis.