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将氨酸转化为基米氨酸:表面循环的作用
L Hedstrom1, L Szilagyi, W J Rutter
1Hormone Research Institute, University of California, San Francisco 94143-0534.
概括
改变试素的改变
科学领域:
- 生物化学 生化学
- 酶学 是一种酶学.
- 蛋白质工程是指蛋白质工程.
背景情况:
- 素和胆素是类似结构但具有不同的基质特异性的血清蛋白酶.
- 素在基本残留物 (Arg,Lys) 之后分裂,而基米素则准大型疏水性残留物.
- 了解这些特异性差异可以为向应用的酶工程提供信息.
研究的目的:
- 调查青素和化学青素基质特异性的结构决定因素.
- 为了设计一种素突变的基质特异性类似于基素的素突变.
- 阐明S1结合部位和表面环在酶催化和区分中的作用.
主要方法:
- 用局部导向的突变发生来用化学素残留物取代素的S1结合部位.
- 进一步的修改包括在辛和化学辛之间交换表面环 (残留物185-188和221-225).
- 在野生类型和突变酶上进行了酶动力学和基质结合试验.
主要成果:
- 仅仅替换S1位点就转移了酶特异性,但没有转移胺解特异性.
- 突变S1位点和特定的表面环都将氨酸转化为一种类似于化学氨酸的酶.
- 改造的突变体表现出甲基胺的催化率,但其基质结合功能受损.
- 突变物体的基质歧视发生在化过程中,而不是结合,类似于基米.
结论:
- 表面环,不直接是活性部位的一部分,显著影响蛋白酶基质的特异性.
- 酶特异性是活性部位和远程结构元素之间的复杂相互作用.
- 这项研究提供了对蛋白质工程的见解,用于创建具有新型催化性能的酶.
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