Arabinose-induction of lac-derived promoter systems for penicillin acylase production in Escherichia coli.
Arabinose was shown to serve as an effective inducer for induction of the lac-derived promoters in Escherichia coli using penicillin acylase (PAC) as a model protein. Upon the induction with a conventional inducer, isopropyl-beta-d-thiogalactopyranoside (IPTG), for pac overexpression, which is regulated by the trc or (DE3)/T7 promoter, the production of PAC was limited by the accumulation of PAC precursors (proPAC) as inclusion bodies. Negative cellular responses, such as growth inhibition and cell lysis, were frequently observed, resulting in a low pac expression level and poor culture performance. Interestingly, these technical hurdles can be overcome simply through the use of arabinose as an inducer. The results indicate that arabinose not only induced the lac-derived promoter systems (i.e., trc and (DE3)/T7) for pac (or LL pac) overexpression but also facilitated the posttranslational processing of proPAC for maturation. However, the arabinose-inducibility appears to be host-dependent and becomes less observable in the strains with a mutation in the ara operon. The arabinose-inducibility was also investigated in the expression system with the coexistence of the trc promoter system regulating pac expression and another arabinose-inducible promoter system of araB regulating degP coexpression.
Narayanan
Niju
N
Department of Chemical Engineering, University of Waterloo, Waterloo, Ontario, Canada N2L 3G1.
Hsieh
Ming-Yi
MY
Xu
Yali
Y
Chou
C Perry
CP
D001089:Arabinose / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000187:drug effects; D002478:Cells, Cultured; D004926:Escherichia coli / Q000187:drug effects* / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects; D008562:Membrane Glycoproteins / Q000096:biosynthesis / Q000187:drug effects; D010399:Penicillin Amidase / Q000096:biosynthesis* / Q000187:drug effects*; D011401:Promoter Regions, Genetic / Q000187:drug effects* / Q000235:genetics / Q000502:physiology*
D001426:Bacterial Proteins; D008562:Membrane Glycoproteins; C055764:S-layer proteins; D001089:Arabinose; D010399:Penicillin Amidase