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通过染色体整合表达古菌乙酰化酶合成N-末端乙酰化胸腺肽β4

DOI: 10.1360/052011-565, PP. 1008-1015

Keywords: 胸腺肽β4,乙酰化酶,N-末端乙酰化,Intein

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Abstract:

胸腺肽β4(Tβ4)是N-末端乙酰化的43肽,具有多种重要生物学功能.其生物合成存在两大难点,即乙酰化修饰和小分子肽的表达.本研究发现来自古菌Sulfolobussolfataricus的乙酰化酶ssArd1可以催化Tβ4的N-末端乙酰化修饰.利用Red同源重组技术将ssArd1基因表达盒整合至E.coliBL21(DE3)染色体的lpxM位点上,构建了可以实现Tβ4N-末端乙酰化修饰的新型宿主E.coliBDA.将Tβ4编码基因融合在改造的微型SplDnaXIntein的N端,并在Intein的C端添加His标签,构建了表达载体pET-Tβ4-Intein.在E.coliBDA中表达的融合蛋白,经镍亲和层析纯化后用β-巯基乙醇诱导融合蛋白切割释放小分子多肽,获得了具有N-末端乙酰化的Tβ4.

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