Carboxyl-terminal domain characterization of polyene-specific P450 hydroxylase in Pseudonocardia autotrophica. (1st November 2016)

• Record Type: Journal Article
• Authors: Kim, Min-Kyung; Won, Hyung-Jin; Kim, Hye-Jin; Choi, Si-Sun; Lee, Heung-Shick; Kim, Pil; Kim, Eung-Soo
• Is Part Of: Journal of industrial microbiology & biotechnology. Volume 43:Number 11(2016)
• Page Start: 1625
• Page End: 1630
• Publication Date: 2016-11-01
• Subjects: Polyene -- P450 hydroxylase -- Pseudonocardia -- Substrate specificity; Industrial microbiology -- Periodicals; 660.62
• DOI: 10.1007/s10295-016-1813-z
• Languages: English
• ISSNs: 1367-5435

Abstract

Abstract: A polyene compound NPP identified in Pseudonocardia autotrophica was shown to contain an aglycone identical to nystatin, but to harbor a unique disaccharide moiety that led to higher solubility and reduced hemolytic activity. Recently, it was revealed that the final step of NPP (nystatin-like polyene) biosynthesis is C10 regio-specific hydroxylation by the cytochrome P450 hydroxylase (CYP) NppL (Kim et al. [7 ]). Through mutation and cross-complementation, here we found that NppL preferred a polyene substrate containing a disaccharide moiety for C10 hydroxylation, while its orthologue NysL involved in nystatin biosynthesis showed no substrate preference toward mono- and disaccharide moieties, suggesting that two homologous polyene CYPs, NppL and NysL might possess a unique domain recognizing a sugar moiety. Two hybrid NppL constructs containing the C-terminal domain of NysL exhibited no substrate preference toward 10-deoxy NPP and 10-deoxy nystatin-like NysL, implying that the C-terminal domain plays a major role in differentiating the sugar moiety responsible for substrate specificity. Further C-terminal domain dissection of NppL revealed that the last fifty amino acids play a critical role in determining substrate specificity of polyene-specific hydroxylation, setting the stage for the biotechnological application of hydroxyl diversification for novel polyene biosynthesis in actinomycetes.