Gene Expression of Lytic Endopeptidases AlpA and AlpB from Lysobacter sp. XL1 in Pseudomonads

Abstract

Development of an efficient expression system for (especially secreted) bacterial lytic enzymes is a complicated task due to the specificity of their action. The substrate for such enzymes is peptidoglycan, the main structural component of bacterial cell walls. For this reason, expression of recombinant lytic proteins is often accompanied with lysis of the producing bacterium. This paper presents data on the construction of an inducible system for expression of the lytic peptidases AlpA and AlpB from <i>Lysobacter</i> sp. XL1 in <i>Pseudomonas fluorescens</i> Q2-87, which provides for the successful secretion of these proteins into the culture liquid. In this system, the endopeptidase gene under control of the T7<i>lac </i>promoter was integrated into the bacterial chromosome, as well as the <i>Escherichia coli</i> lactose operon repressor protein gene. The T7 <i>pol</i> gene under <i>lac </i>promoter control, which encodes the phage T7 RNA polymerase, is maintained in <i>Pseudomonas</i> cells on the plasmids. Media and cultivation conditions for the recombinant strains were selected to enable the production of AlpA and AlpB by a simple purification protocol. Production of recombinant lytic enzymes should contribute to the development of new-generation antimicrobial drugs whose application will not be accompanied by selection of resistant microorganisms.