Overexpression of the pectin lyase gene of Pseudomonas marginalis in Escherichia coli and purification of the active enzyme.

@article{Papi2003OverexpressionOT,
  title={Overexpression of the pectin lyase gene of Pseudomonas marginalis in Escherichia coli and purification of the active enzyme.},
  author={Rigini M. Papi and Dimitrios A. Kyriakidis},
  journal={Biotechnology and applied biochemistry},
  year={2003},
  volume={37 Pt 2},
  pages={187-94}
}
A pectin lyase gene (pnl) of Pseudomonas marginalis was cloned and overexpressed in Escherichia coli BL21(DE3). The pnl gene was amplified by PCR, inserted into pET29c with a six-His tag and the overproduced active enzyme was purified almost to homogeneity using a Ni(2+)-nitrilotriacetate-agarose column. The purified pectin lyase (PNL; EC 4.2.2.10, family 1) is inhibited by NAD+ (at concentrations above 0.25 mM), NADH or dithiothreitol. Evidence for the existence of a heat-labile protein… CONTINUE READING