Open AccessEffect of Signal Peptides on the Secretion of β-Cyclodextrin Glucanotransferase in <b><i>Lactococcus lactis</i></b> NZ9000
Author(s) -
Menaga Subramaniam,
Ali Baradaran,
Illias Rosli,
Rosfarizan Mohamad,
Khatijah Yusoff,
Abdul Rahim Raha
Publication year - 2012
Publication title -
microbial physiology
Language(s) - English
Resource type - Journals
eISSN - 2673-1673
pISSN - 2673-1665
DOI - 10.1159/000343921
Subject(s) - lactococcus lactis , biochemistry , secretion , extracellular , chemistry , plasmid , cyclodextrin , starch , lactic acid , biology , bacteria , dna , genetics
Cyclodextrin glucanotransferase (CGTase) is an extracellular enzyme which catalyzes the formation of cyclodextrin from starch. The production of CGTase using lactic acid bacterium is an attractive alternative and safer strategy to produce CGTase. In this study, we report the construction of genetically modified Lactococcus lactis strains harboring plasmids that secrete the Bacillus sp. G1 β-CGTase, with the aid of the signal peptides (SPs) SPK1, USP45 and native SP (NSP). Three constructed vectors, pNZ:NSP:CGT, pNZ:USP:CGT and pNZ:SPK1:CGT, were developed in this study. Each vector harbored a different SP fused to the CGTase. The formation of halo zones on starch plates indicated the production and secretion of β-CGTase by the recombinants. The expression of this enzyme is shown by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and zymogram analysis. A band size of ∼75 kDa corresponding to β-CGTase is identified in the intracellular and the extracellular environments of the host after medium modification. The replacement of glucose by starch in the medium was shown to induce β-CGTase production in L. lactis. Although β-CGTase production is comparatively low in NZ:SPK1:CGT, the SP SPK1 was shown to have higher secretion efficiency compared to the other SPs used in this study.