Biochemical genetics of Cephalosporin C production
The biosynthetic pathway which leads, in Cephalosporium acremonium, to the production of the commercially important β-lactam antibiotic Cephalosporin C (CPC) has been the subject of extensive biochemical studies and is now well characterized. In contrast, genetic analysis in this organism was limited until the application of protoplast fusion techniques facilitated parasexual analysis and allowed a genetic map to be established. (Hamlyn 1982; Hamlyn et al 1985). Subsequently, work leading to our understanding of the genetic basis of the CPC biosynthetic pathway in C. acremonium began. (Perez-Martinez 1984; Perez-Martinez and Peberdy in preparation). The studies described here were aimed at extending this understanding to a point at which individual genes implicated in the pathway could be identified and positioned on the linkage map. A programme of mutagenesis resulted in the production of a number of 'blocked' mutant strains of C. acremonium which were phenotypically particular steps of the CPC biosynthetic pathway. The segregation of several of these mutations relative to other genetic markers was examined. Crosses designed to detect complementation between mutations resulting in a 'blocked' phenotype were carried out and involved strains produced in other laboratories in addition to those characterized during this work. Complementation was shown between two mutations which apparently affected the same step in CPL biosynthesis (the conversion of penicillin N into deacetoxycephalosporin C) and evidence for the linkage of one of the mutations (cnp-6) to a mutation resulting in a requirement for inositol was obtained. During the course of the complementation studies, it was noted that the haploid and heterozygous products obtained following C. acremonium protoplast fusion crosses did not always behave in the typical manner described previously. (Hamlyn 1984). The persistent heterogeneity of these fusion products and the possible implications of this are discussed.