Impacting of polymorphism of beta-amyrin synthase from Glycyrrhiza uralensis on its catalytic efficiency / 中国中药杂志

ABSTRACT

<p><b>OBJECTIVE</b>To analyze heterologous expression in Saccharomyces cerevisiae of two genotypes beta-AS (A-T) genotype which is related to high content of glycyrrhizic acid and beta-AS(G-C) genotype which is related to low content of glycyrrhizic acid, and compare two different genotypes on the impact of beta-amyrin production in order to provide a foundation for licorice molecular breeding.</p><p><b>METHOD</b>The 2 289 bp fragment in plasmid pMD-19T encoding beta-amyrin synthase was subcloned into the yeast-Escherichia coli shuttle vector pY26, thus an expression recombinant plasmid PY-beta-AS containing target gene was constructed. The PY-beta-AS was introduced into defective mutant INVSc1 of S. cerevisiae by LiAc method, after induced by IPTG, the content of beta-amyrin was determined by GC-MS.</p><p><b>RESULT</b>GC-MS analysis demonstrates that the an occurring peak corresponding to beta-amyrin standards was detected with the same retention time, which is absent in the cell transform with empty vector. Results showed the peak was beta-amyrin and the percentage of beta-amyrin in two genotypes beta-AS (A-T) genotype and beta-AS (G-C) genotype were 19.08% and 1.40%, respectively. Thus the beta-amyrin synthase exhibited the activity of catalyzing 2, 3-oxidosqualene to beta-amyrin.</p><p><b>CONCLUSION</b>The catalytic efficiency of beta-AS(A-T) genotype is higher than that of beta-AS(G-C) genotype, which can lay the foundation for licorice molecular breeding.</p>