The orthosomycins are highly modified oligosaccharide natural products with a broad spectrum and potent antimicrobial activities. These include everninomicins and avilamycins, which inhibit protein translation by binding a unique site on the bacterial ribosome. Notably, ribosomal bound structures reveal a network of interactions between the 50S subunit and dichloroisoeverninic acid (DCIE), the aromatic A-ring conserved across orthosomycins, but the relationship of these interactions to their antimicrobial activity remains undetermined. Genetic functional analysis of three genes putatively associated with DCIE biosynthesis in the everninomicin producer delineates the native biosynthetic pathway and provides previously unreported advanced biosynthetic intermediates. Subsequent biochemical analyses demonstrate the complete DCIE biosynthetic pathway and provide access to novel everninomicin analogs. In addition to the orsellinate synthase EvdD3 and a flavin-dependent halogenase EvdD2, our results identified a key acyltransferase, EvdD1, responsible for transferring orsellinate from the acyl carrier protein domain of EvdD3 to a heptasaccharide orthosomycin biosynthetic intermediate. We have also shown that EvdD1 is able to transfer unnatural aryl groups via their -acyl cysteamine thioesters to the everninomicin scaffold and used this as a biocatalyst to generate a panel of unnatural aryl analogs. The impact of diverse aryl functional group substitution on both ribosome inhibition and antibacterial activities demonstrates the importance of the DCIE moiety in the pharmacology of orthosomycins, notably revealing an uncoupling between ribosomal engagement and antibiotic activity. Control of A-ring functionality in this class of molecules provides a potential handle to explore and address pharmacological roles of the DCIE ring in this potent and unique class of antibiotics.
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http://dx.doi.org/10.1021/acschembio.3c00693 | DOI Listing |
ACS Chem Biol
February 2024
Department of Chemistry, Vanderbilt University, Nashville, Tennessee 37235, United States.
The orthosomycins are highly modified oligosaccharide natural products with a broad spectrum and potent antimicrobial activities. These include everninomicins and avilamycins, which inhibit protein translation by binding a unique site on the bacterial ribosome. Notably, ribosomal bound structures reveal a network of interactions between the 50S subunit and dichloroisoeverninic acid (DCIE), the aromatic A-ring conserved across orthosomycins, but the relationship of these interactions to their antimicrobial activity remains undetermined.
View Article and Find Full Text PDFFood Chem
May 2018
Division of Foods, National Institute of Health Sciences, Tonomachi 3-25-26, Kawasaki-ku, Kawasaki, Kanagawa 210-9501, Japan. Electronic address:
A sensitive and reliable method for determining the total avilamycin residues was developed using LC-MS/MS. Avilamycin (consisting of avilamycin A and 15 other minor factors) and its metabolites in porcine muscle, fat, and liver were analysed as the marker residue dichloroisoeverninic acid (DIA), in accordance with the maximum residue limit (MRL) established by international organisations such as Codex Alimentarius Commission and other regulatory bodies. The analytes were extracted from samples with acetone, hydrolysed to DIA, partitioned into ethyl acetate, and cleaned up prior to the LC-MS/MS analysis.
View Article and Find Full Text PDFAnal Bioanal Chem
October 2013
Analytical and Advisory Services Division, Government Laboratory, 7/F, Ho Man Tin Government Offices, 88, Chung Hau Street, Ho Man Tin, Kowloon, Hong Kong, China.
Avilamycin residue in food is regulated as its marker residue dichloroisoeverninic acid (DIA). An isotope dilution liquid chromatography-tandem mass spectrometry method is established for the accurate determination of DIA in animal muscles without any pre-extraction and preconcentration prior to alkaline hydrolysis. Optimization of the sample cleanup procedures such as liquid-liquid extraction and solid phase extraction was performed by fine-tuning several critical parameters to reduce the matrix effects.
View Article and Find Full Text PDFJ Biol Chem
May 2006
Institut für Pharmazeutische Wissenschaften, Pharmazeutische Biologie und Biotechnologie, Albert-Ludwigs-Universität Freiburg, Stefan-Meier-Strasse 19, D-79104 Freiburg, Germany.
The antibiotic avilamycin A is produced by Streptomyces viridochromogenes Tü57. Avilamycin belongs to the family of orthosomycins with a linear heptasaccharide chain linked to a terminal dichloroisoeverninic acid as aglycone. The gene cluster for avilamycin biosynthesis contains 54 open reading frames.
View Article and Find Full Text PDFChem Biol
October 2005
Institut für Pharmazeutische Wissenschaften, Pharmazeutische Biologie und Biotechnologie, Albert-Ludwigs-Universität Freiburg, Germany.
The oligosaccharide antibiotic avilamycin A is composed of a polyketide-derived dichloroisoeverninic acid moiety attached to a heptasaccharide chain consisting of six hexoses and one unusual pentose moiety. We describe the generation of mutant strains of the avilamycin producer defective in different sugar biosynthetic genes. Inactivation of two genes (aviD and aviE2) resulted in the breakdown of the avilamycin biosynthesis.
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