Effects of Signal Peptide on the Secretory Expression of Multifunctional Amylase OPMA and Its Catalytic Activity

doi:10.7503/cjcu20130337

Abstract

Abstract:

The recombinant expression vectors pMA5-OPMA-SN and pMA5-OPMA-SG were constructed by fusing the gene encoding the signal peptide(S) of amyE from Bacillus subtilis 168 with the gene encoding the multifunctional amylases OPMA-N or OPMA-G, and the high-level secretory expression of OPMA-N and OPMA-G were achieved by Bacillus subtilis 168. SDS-PAGE analysis showed that the introduction of the signal peptide was indeed possible to improve the secretion of heterologous protein, and the secretory yields of OPMA-SN or OPMA-SG(53% or 67%) were obviously higher than those of OPMA-N or OPMA-G(45% or 58%), but the signal peptide could not be effectively removed after secretion. The activity analysis demonstrated that the levels of OPMA-SN and OPMA-N activities(5.17 and 4.57 U/mg) were consistent with their secretory yields, but it was not the case for OPMA-SG(4.50 U/mg) and OPMA-G(4.65 U/mg), there was a negative correlation between the activity levels and secretory yields of for OPMA-SG and OPMA-G. The spatial conformation of OPMA-SN and OPMA-SG molecules proved that the presence of the signal peptide at the N terminus of OPMA-N or OPMA-G could not affect the conformation of the catalytic centre in OPMA-N, but affected that in OPMA-SG and resulted in the decrease of OPMA-SG's catalytic activity. These findings could provide important guidance on signal peptide application for secretory expression of heterologous proteins in Bacillus subtilis, and for the structure and function of the target protein.

Key words: Multifunctional amylase, Expression, Signal peptide, Structure, Function

CLC Number:

TrendMD:

LI Yan-Fei, ZHANG Jin-Xiang, GU Yan-Qin, HAO Qian, CAO Hao, ZHANG Ying-Jiu. Effects of Signal Peptide on the Secretory Expression of Multifunctional Amylase OPMA and Its Catalytic Activity[J]. Chem. J. Chinese Universities, 2013, 34(10): 2334.

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