title: Carboxyethylarginine synthase genes show complex cross-regulation in Streptomyces clavuligerus.
abstract: Carboxyethylarginine synthase is the first dedicated enzyme of clavam biosynthesis in Streptomyces clavuligerus and is present in two isoforms encoded by two separate genes. When grown on a liquid soy medium, strains with ceaS1 deleted showed only a mild reduction of clavam biosynthesis, while disruption of ceaS2 abolished all clavam biosynthesis. Creation of an in-frame ceaS2 deletion mutant to avoid polarity did not restore clavam production, nor did creation of a site-directed mutant altered only in a single amino acid residue important for activity. Reverse transcriptase PCR analyses of these mutants indicated that the failure to produce clavam metabolites could be traced to reduced or abolished transcription of ceaS1 in the ceaS2 mutants, despite the location of ceaS1 on a replicon completely separate from that of ceaS2. Western analyses further showed that the CeaS1 protein (as well as the CeaS2 protein) was absent from the ceaS2 mutants. Complementation experiments were able to restore clavam production partially, but only by virtue of restoring CeaS2 production. CeaS1 was still absent from the complemented strains. While this dependence of CeaS1 production on the expression of ceaS2 from its native chromosomal location was seen in all of the ceaS2 mutants, the effect was limited to growth in liquid medium. When the same mutants were grown on solid soy medium, clavam production was restored and CeaS1 was produced, albeit at low levels compared to the wild type.
authors.affiliation: Department of Biological Sciences, University of Alberta, Edmonton, Alberta, Canada.
authors.forename: Thomas
authors.initials: T
authors.lastname: Kwong
authors.forename: Kapil
authors.initials: K
authors.lastname: Tahlan
authors.forename: Cecilia L
authors.initials: CL
authors.lastname: Anders
authors.forename: Susan E
authors.initials: SE
authors.lastname: Jensen
mesh_terms: D019943:Amino Acid Substitution; D015153:Blotting, Western; D002969:Clavulanic Acids / Q000096:biosynthesis*; D003470:Culture Media / Q000737:chemistry; D017353:Gene Deletion; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial; D005816:Genetic Complementation Test; D013302:Streptomyces / Q000201:enzymology* / Q000235:genetics*
chemical_list: D002969:Clavulanic Acids; D003470:Culture Media; C483963:clavam