Detection and Expression of Biosynthetic Genes in Actinobacteria ...

BERVANAKIS, G.Chapter 1: INTRODUCTIONsequence identities between Streptomycete spp. (Aharonowitz et al., 1992). The cefEgene encodes the enzyme deacetoxycephalosporin C synthase (DAOCS). TheDAOCS enzyme catalyzes the oxidation reaction converting the five-memberedthiazolidine ring of penicillin into six-membered dihydrothiazine ring [Figure 13; step4].. The cefF gene encodes the enzyme deacetylcephalosporin C hydroxylase whichperforms a hydroxylation reaction forming deacetylcephalosporin C (DAC) [Figure13; step 5].In this study the pcbC gene was targeted. The high DNA and protein sequencehomologies of this gene consists of between 79 – 85 % nucleotide and 58 – 64 %amino acid sequence identities. The high nuceleotide sequence homologies hasenabled the efficient isolation of the pcbC gene in β-lactam producing actinobacteria.(Aharonowitz et al., 1992; Sim & Loke, 2000). Other β-lactam biosynthetic genes thatcould have been screened include pcbAB gene which contains 48 % nucleotidesequence identity between actinobacteria producing β-lactam antibiotics.Section 4: Gene Expression in Actinobacteria1.4.1 Regulation of Gene Expression in ActinobacteriaThe biosynthesis of certain secondary metabolites are regulated by specific sets ofstructural genes. In general, these genes are clustered on the chromosome in the orderin which the biosynthetic reaction will proceed. Regulatory genes (listed in Table 6)are juxtaposed to these biosynthetic genes which control specific SM biosyntheticpathways. The expressivity of genes to changing environmental conditions, eg.nutritional imbalance, leads to the activation or suppression of secondary metabolitebiosynthetic genes [SMBG] (van Wezel et al., 1997; Narberhaus, 1999; Vicente et al.,1999). Pathway-specific regulators listed in table 6 play crucial roles in controllingnumerous secondary metabolite biosynthetic genes. Figure 14 depicts the regulatoryevents leading to the formation of a SM in a bacterial cell._____________________________________________________________________24