Engineering of an intersubunit disulfide bridge in the iron-superoxide dismutase of Mycobacterium tuberculosis
Engineering of an intersubunit disulfide bridge in the iron-superoxide dismutase of Mycobacterium tuberculosis
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DOI：
10.1006/abbi.2001.2635
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发表时间：
2002-01-01
影响因子：
3.9
通讯作者：
Young, DB
中科院分区：
文献类型：
--
作者：
Bunting, KA;Cooper, JB;Young, DB
为了加强参与二聚体形成的相互作用，在结核分枝杆菌的超氧化物歧化酶中设计了亚基间二硫化物桥。选择Ser-123突变为半胱氨酸是因为它位于二聚体界面，丝氨酸侧链与相反亚基的相同残基相互作用。表达突变体的凝胶电泳和x射线晶体学研究证实在非还原条件下形成了二硫键。然而，根据在盐酸胍存在下对变性的敏感性判断，发现突变蛋白比野生型更不稳定。稳定性下降可能是由于形成了一个具有次优扭转角的二硫桥，并且溶剂分子被排除在二聚体界面之外。(C) 2001 Elsevier Science。
With the aim of enhancing interactions involved in dimer formation, an intersubunit disulfide bridge was engineered in the superoxide dismutase enzyme of Mycobacterium tuberculosis. Ser-123 was chosen for mutation to cysteine since it resides at the dimer interface where the serine side chain interacts with the same residue in the opposite subunit. Gel electrophoresis and X-ray crystallographic studies of the expressed mutant confirmed formation of the disulfide bond under nonreducing conditions. However, the mutant protein was found to be less stable than the wild type as judged by susceptibility to denaturation in the presence of guanidine hydrochloride. Decreased stability probably results from formation of a disulfide bridge with a suboptimal torsion angle and exclusion of solvent molecules from the dimer interface. (C) 2001 Elsevier Science.