390. Characterization of the Penicillium paf gene promoter for gene expression in Penicillum chrysogenum. R. Kerkman, A.W.H. Vollebregt, R.A.L. Bovenberg. Gist-brocades, IPPD/PDD, Delft, The Netherlands.
In contrast to Aspergillus, only few Penicillium promoters have been characterised so far. In order to isolate and analyse strong promoters for gene expression in Penicillium we examined the relative strength of the promoters of the Penicillium pcbC and paf genes in P. chrysogenum. The promotor of pcbC, is one of most extensively investigated promoters and is considered to be a strong promotor. The paf gene, which encodes a 12-kDa protein with potential antifungal activity, is one of the most abundant proteins present in the culture medium (Vollebregt, ECFG2 abstract B48). To analyse the paf gene promotor in more detail integration vectors were constructed containing various parts/fragments of the promotor region fused to the Streptomyces clavuligerus cefE gene encoding expandase which can be used as a reporter gene. In the presence of suitable side chains e.g. adipate Penicillium transformants expressing expandase are able to convert penicillins to cephalosporins, which can be measured by using bio-assays and shake flask fermentations.Transformants were obtained by co-transformation using the Aspergillus amdS gene as a selectable marker. Selected transformants were tested for promoter activity by performing shake flask experiments and measuring formation of adipoyl- 7-ADCA. The results which will be presented indicate that the paf gene promotor is a strong/suitable promotor for heterologous gene expression in P. chrysogenum.