Abbruzzetti S, Viappiani C, Bruno S, Bettati S, Bonaccio M, Mozzarelli A
National Institute for the Physics of Matter (INFM), Department of Physics, University of Parma, Via Parco delle Scienze, 43100 Parma, Italy.
J Nanosci Nanotechnol. 2001 Dec;1(4):407-15. doi: 10.1166/jnn.2001.058.
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.
肌红蛋白和血红蛋白被封装在湿的纳米多孔硅胶中。通过纳秒激光闪光光解后用瞬态吸收检测测量一氧化碳重新结合的动力学，以及用吸收显微分光光度法测量氧亲和力，对封装对蛋白质动力学和功能的影响进行了严格评估。一氧化碳与肌红蛋白结合的时间过程表明，相对于溶液，双分子复合显著增强且双分子重新结合更快，而血红蛋白的T态和R态四级结构表现出与溶液中观察到的非常相似的双分子复合阶段和双分子结合速率。发现T态血红蛋白的氧亲和力与溶液中T态血红蛋白结合第一个氧时观察到的氧亲和力相近。结果表明，一些构象转变在动力学上受到限制，这使得能够分离出不同的三级和四级结构状态。这为其详细的功能表征及其在生物器件中的应用开辟了道路。
