Strategy for successful expression of the Pseudomonas putida nitrile hydratase activator P14K in Escherichia coli.

ABSTRACT:

Background

Activators of Nitrile hydratase (NHase) are essential for functional NHase biosynthesis. However, the activator P14K in P. putida is difficult to heterogeneously express, which retards the clarification of the mechanism of P14K involved in the maturation of NHase. Although a strep tag containing P14K (strep-P14K) was over-expressed, its low expression level and low stability affect the further analysis.

Results

We successfully expressed P14K through genetic modifications according to N-end rule and analyzed the mechanism for its difficult expression. We found that mutation of the second N-terminal amino-acid of the protein from lysine to alanine or truncating the N-terminal 16 amino-acid sequence resulted in successful expression of P14K. Moreover, fusion of a pel

SUBMITTER: Liu Y 

PROVIDER: S-EPMC3680314 | biostudies-literature | 2013 Jun

REPOSITORIES: biostudies-literature