Proudfoot A E, Wallace C J
Université de Genève, Département de Biochimie Médicale, Switzerland.
Biochem J. 1987 Dec 15;248(3):965-7. doi: 10.1042/bj2480965.
We have used chemical and enzymic protein engineering techniques to create analogues of the semisynthetic two-fragment complex (1-37).(38-104) of mitochondrial cytochrome c. This complex, unlike the natural product of specific tryptic cleavage, (1-38).(39-104), from which it is prepared, quite closely resembles the parent protein in functional characteristics and is thus a suitable substrate for modifications designed to study structure-function relations. We have replaced the invariant Arg-38 and the conserved Lys-39 with a range of alternative amino acids and have studied the effects on the principal functional parameters. The hydrogen-bonding capacity of Arg-38 is crucial to the stabilization of the bottom omega-loop, while the positive charge of Lys-39 helps maintain the high redox potential by electrostatic effects at the haem iron.
我们运用化学和酶促蛋白质工程技术,构建了线粒体细胞色素c半合成双片段复合物(1 - 37).(38 - 104)的类似物。该复合物与用于制备它的特定胰蛋白酶切割天然产物(1 - 38).(39 - 104)不同,在功能特性上与亲本蛋白非常相似,因此是用于研究结构 - 功能关系的修饰的合适底物。我们用一系列替代氨基酸取代了不变的精氨酸 - 38和保守的赖氨酸 - 39,并研究了其对主要功能参数的影响。精氨酸 - 38的氢键结合能力对于底部ω - 环的稳定至关重要,而赖氨酸 - 39的正电荷通过对血红素铁的静电作用有助于维持高氧化还原电位。