Burkholderia cepacia XYU-6脂肪酶的克隆及其细胞表面展示

通过分析GenBank中Burkholderia cepacia脂肪酶的序列，设计简并引物，采用同源克隆的策略，成功地从B. cepacia XYU-6菌株中克隆到脂肪酶基因bcl，其大小为1095 bp，编码364个氨基酸（ GenBank登陆号KR233260）.将bcl基因与质粒pET-28b（＋）连接并转化大肠杆菌，使脂肪酶BCL的大肠杆菌胞内过表达，其活力是野生菌的12.9倍.通过将bcl基因克隆到细胞表面展示载体pZXL中，构建脂肪酶BCL的细胞表面展示工程菌，使BCL在Lpp-OmpA引导下定位于大肠杆菌细胞表面，其活力是野生菌的3.9倍.研究结果为脂肪酶BCL后续的分子改造和应用奠定基础.

The Cloning and Cell Surface Display of a Lipase from Burkholderia cepacia XYU-6

The lipase gene bcl was isolated from Burkholderia cepacia XYU-6 by homologous cloning method using degenerate primers,which was design based on the analyzing the sequences of lipases from B. cepacia. The gene bcl contained a 1 095 bp open reading frame encoding a protein of 364 amino acids( GenBank accession number:KR233260 ). The gene bcl was cloned into the expression vector pET-28 b( +),and intracellular overexpressed in biologically active in Escherichia coli. Meanwhile,the gene bcl was cloned into the cell-surface display vector pZXL and transformed into E. coli. The lipase BCL was successfully achieved on the cell surface of engineering strain using the anchoring motif Lpp-OmpA. The two recombinant strains demonstrated 12. 9-fold and 3. 9-fold of lipase activity compared to the wild B. cepacia XYU-6,respectively. This study paves the way for the further research of the lipase for protein engineering and application in the industry.