The use of specialised transducing phages in the amplification of enzyme production

Abstract

Two types of lambdatrp phages have been used as model systems to investigate ways of optimising the expression of bacterial genes from transducing phage genomes. Excellent yields of trp enzymes were achieved by infecting a trpR- host with Q- or Q-S- derivatives of lambdatrpAM1, which expresses its trp genese exclusively from the trp promoter. The five trp geneproducts constituted more than 50% of the total soluble protein of infected cells under these conditions, and an even higher proportion of the protein synthesized after infection. In a trpR+ host, phage DNA replication was easily able to override tryptophan-mediated repression by titration of the trp promoter were equally productive, while having the advantage of being much simpler to construct and propagate. lambdatrp phages lacking the trp promoter were used to investigate ways of optimising gene expression initiated at the phage promoter, PL. Though very powerful, the latter promoter is more difficult to harness then the trp promoter. Derepression of transcription from PL by the use of cro- mutations is accompanied by poor replication of transducing phage DNA. Attempts to circumvent this difficulty using virulent of cro,cII double mutants have not been successful. Nevertheless, cells infected with a lambdatrp phage expressing its trp genes exclusively from PL made up to 16 per cent of their protein as trp gene-products.