利用非经典分泌蛋白质实现脂肪酶A的分泌表达

Effect of non-classical secreted proteins on LipaseA secretion

摘要:

摘要:【目的】以枯草芽孢杆菌脂肪酶A(LipaseA)为报告蛋白，尝试利用4种非经典分泌蛋白质及其前50个氨基酸作为分泌信号以实现其分泌表达。【方法】我们扩增了脂肪酶A 的编码基因和非经典分泌蛋白质的编码序列，构建了8种针对脂肪酶A的分泌表达载体，并转化至枯草芽孢杆菌WB800菌株，通过测定重组菌株的酶活、利用蛋白质电泳和免疫印迹等技术检测脂肪酶A的分泌情况【结果】以PdhA的氨基酸序列和SodA、Eno的前50氨基酸序列作为分泌信号的重组菌株较好的实现了脂肪酶A的分泌表达。【结论】部分非经分泌蛋白质的编码基因或其前50个氨基酸序列能够引导脂肪酶A分泌至细胞外。

关键词:关键词:枯草芽孢杆菌，非经典分泌蛋白质，脂肪酶A

Abstract:

Abstract:［Objective］We used 50-amino acid-long peptides from the N-terminus of 4 different non-classically secreted proteins to study the secretion efficiency of Bacillus subtilis LipaseA via non-classical secretion pathway.［Methods］We amplified the coding sequences (CDs) of LipaseA and N-terminus of non-classically secreted proteins，constructed 8 fusion protein expression vectors containing both LipaseA CD and different secretion signal peptide and transformed them into B．subtilis WB800．Secretion efficiency of these fusion proteins was analyzed by enzyme activity，SDS-PAGE and Western-Blot． ［Results］Recombinant LipaseA containing coding sequences of PdhA or N-terminus of SodA and Eno as secretion signals was efficiently secreted．［Conclusion］ Parts of non-classically secreted proteins or N-terminus (50amino acids) could guide LipaseA protein secretion．

Key words:Keywords: Bacillus subtilis，non-classical secreted proteins，LipaseA