HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) Supplementary data Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via CrossRef Citing Articles via Google Scholar Google Scholar Articles by Neary, J. M. Articles by Micklefield, J. Search for Related Content PubMed PubMed Citation Articles by Neary, J. M. Articles by Micklefield, J. Agricola Articles by Neary, J. M. Articles by Micklefield, J.

An asparagine oxygenase (AsnO) and a 3-hydroxyasparaginyl phosphotransferase (HasP) are involved in the biosynthesis of calcium-dependent lipopeptide antibiotics

Colin P. Smith^2,

Jason Micklefield^1,

¹ School of Chemistry, The University of Manchester, PO Box 88, Manchester M60 1QD, UK
² Department of Biomolecular Sciences, UMIST, PO Box 88, Manchester M60 1QD, UK
³ Biotica Technology Ltd, Chesterford Research Park, Little Chesterford, Saffron Walden, Essex CB10 1XL, UK

Correspondence
Jason Micklefield
jason.micklefield{at}manchester.ac.uk

Nonribosomal peptides contain a wide range of unusual non-proteinogenic amino acid residues. As a result, these complex natural products are amongst the most structurally diverse secondary metabolites in nature, and possess a broad spectrum of biological activities. -Hydroxylation of amino acid precursors or peptidyl residues and their subsequent processing by downstream tailoring enzymes are some of the most common themes in the biosynthetic diversification of these therapeutically important peptides. Identification and characterization of the biosynthetic intermediates and enzymes involved in these processes are thus pivotal in understanding nonribosomal peptide assembly and modification. To this end, the putative asparaginyl oxygenase- and 3-hydroxyasparaginyl phosphotransferase-encoding genes hasP and asnO were separately deleted from the calcium-dependent antibiotic (CDA) biosynthetic gene cluster of Streptomyces coelicolor. Whilst the parent strains produce a number of 3-hydroxyasparagine- and 3-phosphohydroxyasparagine-containing CDAs, the hasP mutants produce exclusively non-phosphorylated CDAs. On the other hand, asnO mutants produce several new Asn-containing CDAs not present in the wild-type, which retain calcium-dependent antimicrobial activity. This confirms that AsnO and HasP are required for the -hydroxylation and phosphorylation of the Asn residue within CDA.

Tables detailing the bacterial strains, plasmids, primers and PCR products used in this work, and also a figure showing the PCR analysis of asnO and hasP mutants, are available as supplementary data with the online version of this paper.

Present address: School of Biomedical and Molecular Sciences, University of Surrey, Guildford GU2 7XH, UK.

Present address: Manchester Interdisciplinary Biocentre, The University of Manchester, 131 Princess Street, Manchester M1 7DN, UK.