条形柄锈菌小麦专化型脂肪酶基因PsLIP1的酶学性质分析

脂肪作为条形柄锈菌小麦专化型夏孢子萌发及侵染初期的主要营养物质,酶解产物通过乙醛酸循环转化为糖类成分供病菌生长发育所需。脂肪酶是脂肪代谢的关键酶。基于已测序的基因组序列,利用RT-PCR方法克隆了该病菌脂肪酶基因PsLIP1序列（1 302bp）,编码433个氨基酸的蛋白。在毕赤酵母细胞（GS115）中成功表达PsLIP1后,酶学活性分析显示其最适pH为8.0,最适温度为60℃,此外,发现Zn²⁺、Cu²⁺和Ca²⁺对脂肪酶PsLIP1活力有一定激活作用,Fe²⁺、Mn²⁺则对酶活力有抑制作用。研究结果为进一步解析脂肪酶作用机理奠定基础,同时也为从能量代谢角度制定小麦条锈病防控策略提供理论依据。

Expression and enzymatic characteristics of a lipase gene Ps LIP1 from Puccinia striiformis f. sp. tritici

Fat is the main nutrient in the early stage of germination and infection of Puccinia striiformis f.sp.tritici(Pst)urediniospores.The enzymatic hydrolysate of fat is transformed into sugar through glyoxylate cycle for growth and development of the pathogen.Lipase is the key enzyme of fat metabolism.Based on the published genome sequence,the lipase gene Ps LIP1(1302 bp)from Pst was cloned using RT-PCR,and 433 amino acids were encoded.Ps LIP1 was successfully transformed into Pichia pastoris cells(GS115)and expressed with high activity.Enzyme activity analysis showed that the optimum pH was 8.0 and the optimum temperature was 60°C.Zn2+,Cu2+and Ca2+promoted the lipase activity of PsLIP1,while Fe2+and Mn2+inhibited the activity.The results provide a foundation for further analysis of the mechanism of lipase action and formulation of control strategies of wheat stripe rust based on perspective of energy metabolism.