The reaction of hydroxylamine with bacteriorhodopsin studied with mutants that have altered photocycles: selective reactivity of different photointermediates.
The reaction of hydroxylamine with bacteriorhodopsin studied with mutants that have altered photocycles: selective reactivity of different photointermediates.
复制标题
使用改变光循环的突变体研究羟胺与细菌视紫红质的反应：不同光中间体的选择性反应性。
DOI：
10.1073/pnas.88.6.2583
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发表时间：
1991
影响因子：
11.1
通讯作者：
Khorana,HG
中科院分区：
文献类型：
--
作者：
Subramaniam,S;Marti,T;Rösselet,SJ;Rothschild,KJ;Khorana,HG
细菌视紫红质（bR）中视黄醛希夫碱与水溶性试剂羟胺的反应在光照下增强了2个数量级以上。我们利用这个反应来探测野生型bR和质子传输缺陷突变体在光循环过程中希夫碱反应性的变化。我们在这里报道，在pH 6的光照下，与野生型bR相比，D85N突变体与羟胺的依赖反应速率低20倍，D212N突变体与羟胺的依赖反应速率高4倍以上。野生型bR与D96N和T46V突变体的反应性相似。以前的研究表明，D96N和T46V的替代对M的形成动力学没有显著影响，但对M的衰变速率有显著影响。因此，我们得出结论，羟胺反应发生在M中间体形成之前。它最有可能发生在循环的“L”阶段，反映了由于蛋白质构象的光驱动变化而增加的水对希夫碱的可及性。
The reaction of the retinylidene Schiff base in bacteriorhodopsin (bR) to the water-soluble reagent hydroxylamine is enhanced by greater than 2 orders of magnitude under illumination. We have used this reaction as a probe for changes in Schiff base reactivity during the photocycle of wild-type bR and mutants defective in proton transport. We report here that under illumination at pH 6, the D85N mutant has a 20-fold lower rate and the D212N mutant has a greater than 4-fold higher rate for the light-dependent reaction with hydroxylamine compared with wild-type bR. In contrast, the reactivities of wild-type bR and the D96N and T46V mutants are similar. It has been previously shown that the D96N and T46V replacements have no significant effect on the kinetics of "M" formation but have dramatic effects on rate of the decay of M. We therefore conclude that the hydroxylamine reaction occurs before formation of the M intermediate. Most likely it occurs at the "L" stage of the cycle and reflects increased water accessibility to the Schiff base due to a light-driven change in protein conformation.