Sugimoto M, Oikawa T, Esaki N, Tanaka H, Soda K
Institute for Chemical Research, Kyoto University.
J Biochem. 1988 Dec;104(6):924-6. doi: 10.1093/oxfordjournals.jbchem.a122584.
The gene coding for the Neurospora crassa copper metallothionein (MT) was synthesized and inserted in the lacZ' gene of pUC18 plasmid to give the same translational reading frame as the latter gene. The MT-beta-galactosidase fused gene was expressed in Escherichia coli to produce a fused protein in which the amino and carboxy termini of MT are linked to the beta-galactosidase through methionine residues. An MT derivative containing an extra homoserine residue at the carboxy terminus was prepared by cyanogen bromide cleavage of the fused protein followed by a reverse-phase HPLC separation. The spectral features of the MT derivative and its copper complex were similar to those of the corresponding native MTs.
编码粗糙脉孢菌铜金属硫蛋白（MT）的基因被合成并插入到pUC18质粒的lacZ'基因中，以使其与后一个基因具有相同的翻译阅读框。MT-β-半乳糖苷酶融合基因在大肠杆菌中表达，产生一种融合蛋白，其中MT的氨基和羧基末端通过甲硫氨酸残基与β-半乳糖苷酶相连。通过对融合蛋白进行溴化氰裂解，然后进行反相高效液相色谱分离，制备了一种在羧基末端含有一个额外高丝氨酸残基的MT衍生物。该MT衍生物及其铜配合物的光谱特征与相应的天然MT相似。
