title: Genetic evidence for the involvement of glycosyltransferase PdmQ and PdmS in biosynthesis of pradimicin from Actinomadura hibisca.
abstract: Pradimicins are potent antifungal antibiotics with effective inhibitory effects against HIV-1. Pradimicin A consists of an unusual dihydrobenzo[α]naphthacenequinone aglycone substituted with a combination of D-alanine and two sugar moieties. Detailed genetic studies revealed most steps in pradimicin A biosynthesis, but the glycosylation mechanism remained inconclusive. The biosynthetic gene cluster of pradimicin A contains two putative glycosyltransferases, pdmQ and pdmS. However, the exact involvement of each gene in biosynthesis and the particular steps required for precise structural modification was unknown. In this study, the exact role of each gene was evaluated by insertional inactivation and complementation studies. Analysis of the metabolite from both of the disruption mutants revealed abolishment of pradimicin A and complementation resulted in the recovery of production. After deletion of pdmQ, pradimicin B was found to accumulate, whereas deletion of pdmS resulted in the accumulation of aglycone of pradimicin. Together, these results suggest that pdmS is responsible for the attachment of thomosamine to form pradimicin B which in turn is glycosylated by pdmQ to form pradimicin A. These results allowed us to deduce the exact order of terminal tailoring by glycosylation and provided insight into the mechanism of pradimicin A biosynthesis.
authors.affiliation: Institute of Biomolecule Reconstruction (iBR), Department of Pharmaceutical Engineering, Sun Moon University, #100, Kalsan-ri, Tangjeong-myeon, Asan, Chungnam 333-708, Republic of Korea.
authors.forename: Amit Kumar
authors.initials: AK
authors.lastname: Jha
authors.affiliation: Institute of Biomolecule Reconstruction (iBR), Department of Pharmaceutical Engineering, Sun Moon University, #100, Kalsan-ri, Tangjeong-myeon, Asan, Chungnam 333-708, Republic of Korea.
authors.forename: Sarita
authors.initials: S
authors.lastname: Paudel
authors.affiliation: Institute of Biomolecule Reconstruction (iBR), Department of Pharmaceutical Engineering, Sun Moon University, #100, Kalsan-ri, Tangjeong-myeon, Asan, Chungnam 333-708, Republic of Korea.
authors.forename: Dipesh
authors.initials: D
authors.lastname: Dhakal
authors.affiliation: Institute of Biomolecule Reconstruction (iBR), Department of Pharmaceutical Engineering, Sun Moon University, #100, Kalsan-ri, Tangjeong-myeon, Asan, Chungnam 333-708, Republic of Korea.
authors.forename: Pham Thi Thuy
authors.initials: PT
authors.lastname: Van
authors.affiliation: Institute of Biomolecule Reconstruction (iBR), Department of Pharmaceutical Engineering, Sun Moon University, #100, Kalsan-ri, Tangjeong-myeon, Asan, Chungnam 333-708, Republic of Korea.
authors.forename: Gopal Prasad
authors.initials: GP
authors.lastname: Ghimire
authors.affiliation: Institute of Biomolecule Reconstruction (iBR), Department of Pharmaceutical Engineering, Sun Moon University, #100, Kalsan-ri, Tangjeong-myeon, Asan, Chungnam 333-708, Republic of Korea. Electronic address: sohng@sunmoon.ac.kr.
authors.forename: Jae Kyung
authors.initials: JK
authors.lastname: Sohng
mesh_terms: D039903:Actinobacteria / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism*; D018943:Anthracyclines / Q000378:metabolism*; D000890:Anti-Infective Agents / Q000378:metabolism*; D053898:Biosynthetic Pathways / Q000235:genetics*; D017353:Gene Deletion; D005816:Genetic Complementation Test; D006031:Glycosylation; D016695:Glycosyltransferases / Q000235:genetics / Q000378:metabolism*; D016254:Mutagenesis, Insertional; D000095643:Pradimicins and Benanomicins
chemical_list: D018943:Anthracyclines; D000890:Anti-Infective Agents; D016695:Glycosyltransferases; C058036:pradimicin A; D000095643:Pradimicins and Benanomicins; C058037:pradimicin B
keywords: Actinomadura hibisca; Disruption; Glycosyltransferases; Homologous recombination