title: Engineering the expression and biochemical characteristics of recombinant Candida rugosa LIP2 lipase by removing the additional N-terminal peptide and regional codon optimization.
abstract: Candida rugosa lipase (CRL), an important industrial enzyme, has been established, containing several different isoforms which were encoded by the high-identity lip gene family (lip1 to lip7). In this study, we compared the expression and biochemical characterization with three different engineered lip2 constructions in the yeast Pichia pastoris. Our results showed that lip2 (lip2) has an overall improvement of 50% higher production yield (1.446 U/mL) relative to that of nflip2 (0.964 U/mL) at 7 days of cultivation time. Codon-optimized lip2 (colip2) has a 2.3-fold higher production yield (2.182 U/mL) compared to that of lip2 (noncodon-optimized; 1.446 U/mL) and nflip2 (0.964 U/mL), with a cultivation time of 5 days. This finding demonstrated that the removal of the N-terminus and the regional codon optimization of the lip2 gene fragment at the 5' end can greatly increase the expression level of recombinant LIP2 in the P. pastoris system. The distinct biochemical properties of our purified recombinant nfLIP2 and LIP2 suggested that they are potentially useful for various industrial applications.
authors.affiliation: Department of Food Science and Biotechnology, National Chung Hsing University, Taichung 402, Taiwan. swchang@mail.dyu.edu.tw
authors.forename: Shu-Wei
authors.initials: SW
authors.lastname: Chang
authors.forename: Chia-Fang
authors.initials: CF
authors.lastname: Li
authors.forename: Guan-Chiun
authors.initials: GC
authors.lastname: Lee
authors.forename: Tzunuan
authors.initials: T
authors.lastname: Yeh
authors.forename: Jei-Fu
authors.initials: JF
authors.lastname: Shaw
mesh_terms: D020816:Amino Acid Motifs; D002175:Candida / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D003062:Codon; D005656:Fungal Proteins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D015870:Gene Expression; D008049:Lipase / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D015202:Protein Engineering; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D003062:Codon; D005656:Fungal Proteins; D011994:Recombinant Proteins; D008049:Lipase