Functional characterization of heme proteins encapsulated in wet nanoporous silica gels
Functional characterization of heme proteins encapsulated in wet nanoporous silica gels
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DOI：
10.1166/jnn.2001.058
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发表时间：
2001-12-01
影响因子：
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通讯作者：
Mozzarelli, A
中科院分区：
文献类型：
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作者：
Abbruzzetti, S;Viappiani, C;Mozzarelli, A
肌红蛋白和血红蛋白被包裹在湿的纳米多孔硅胶中。采用瞬态吸收法测定纳秒激光闪光光解后一氧化碳重结合动力学，用吸收显微分光光度法测定氧亲和力，对包封对蛋白质动力学和功能的影响进行了严格的评价。一氧化碳与肌红蛋白结合的时间过程表明，与溶液相比，一氧化碳与肌红蛋白的双相结合增强，双分子结合速度更快，而血红蛋白的T和R季态表现出双相结合，双分子结合率与溶液中观察到的非常相似。发现t态血红蛋白的氧亲和力与溶液中第一个氧与t态血红蛋白的结合接近。结果表明，一些构象转变受到动力学限制，允许分离不同的叔态和第四态。这为它们详细的功能表征和在生物器件中的应用开辟了道路。
Myoglobin and hemoglobin were encapsulated in wet, nanoporous silica gels. A rigorous evaluation of the effect of the encapsulation on protein dynamics and function was carried out by measuring the kinetics of carbon monoxide rebinding after nanosecond laser flash-photolysis with transient absorption detection, and the oxygen affinity with absorption microspectrophotometry. The time course of carbon monoxide binding to myoglobin evidenced a strongly enhanced geminate recombination and a faster bimolecular rebinding with respect to solution, whereas T and R quaternary states of hemoglobin exhibited a geminate phase and a bimolecular binding rate very similar to those observed in solution. Oxygen affinity of T-state hemoglobin was found to be close to that observed for the binding of the first oxygen to T-state hemoglobin in solution. Results indicate that some conformational transitions are kinetically restricted, allowing to isolate distinct tertiary and quaternary states. This opens the way to their detailed functional characterization and application to biodevices.