BioChem - DOS

Combined strategies for improving the heterologous expression of an alkaline lipase from Acinetobacter radioresistens CMC-1 in Pichia pastoris

Process biochemistry

Zhao, X.; Xie, W.; Lin, Y.; Lin, X.; Zheng, S.; Han, S.

In this study, a series of strategies was developed to enhance the expression of an alkaline lipase from Acinetobacter radioresistens (ARL) in Pichia pastoris. Activity of the lipase from recombinant strain carrying a single copy of codon-optimized ARL gene was 65U/mL in shake flask culture with p-nitrophenyl caprylate as the substrate. The lipase yield was increased to 104U/mL by introducing a short N-extension spacer peptide coding for the 10 amino acids (EEAEAEAEPK) between &alpha;-factor signal peptide and ARL. The N-terminal extension spacer did not affect the pH or temperature properties of the recombinant ARL. After the multi-copy constructs were identified by Q-PCR assay, a higher lipase activity of 180U/mL was obtained. Further introduction of the spliced HAC1 gene into multi-copy integrants (>6 copies) extensively enhanced the ARL yield by 30-40%. As a result, the ARL yield reached 1.06x10<SUP>4</SUP>U/mL in a 10-L scaled-up fed-batch fermenter as well as the lipase showed some better properties compared to that wild one from A. radioresistens.

2013

Elsevier Applied Science

1359-5113

48

9

pp.1317-1323

10.1016/j.procbio.2013.06.021

http://click.ndsl.kr/servlet/OpenAPIDetailView?keyValue=05787966&target=NART&cn=NART66566752

Acinetobacter radioresistens CMC-1; ARL; Lipase; Secretion expression; Pichia pastoris