Instituto de Química de São Carlos, Universidade de São Paulo, Sao Paulo, SP, Brazil.
Arch Biochem Biophys. 2012 Mar 1;519(1):46-58. doi: 10.1016/j.abb.2012.01.007. Epub 2012 Jan 24.
The stability of the Glossoscolex paulistus hemoglobin (HbGp), in two iron oxidation states (and three forms), as monitored by optical absorption, fluorescence emission and circular dichroism (CD) spectroscopies, in the presence of the chaotropic agent urea, is studied. HbGp oligomeric dissociation, denaturation and iron oxidation are observed. CD data show that the cyanomet-HbGp is more stable than the oxy-form. Oxy- and cyanomet-HbGp show good fits on the basis of a two state model with critical urea concentrations at 220-222 nm of 5.1±0.2 and 6.1±0.1 mol/L, respectively. The three-state model was able to reveal a subtle second transition at lower urea concentration (1.0-2.0 mol/L) associated to partial oligomeric dissociation. The intermediate state for oxy- and cyanomet-HbGp is very similar to the native state. For met-HbGp, a different equilibrium, in the presence of urea, is observed. A sharp transition at 1.95±0.05 mol/L of denaturant is observed, associated to oligomeric dissociation and hemichrome formation. In this case, analysis by a three-state model reveals the great similarity between the intermediate and the unfolded states. Analysis of spectroscopic data, by two-state and three-state models, reveals consistency of obtained thermodynamic parameters for HbGp urea denaturation.
本文研究了在变性剂脲存在的情况下，通过光学吸收、荧光发射和圆二色性（CD）光谱监测，处于两种铁氧化态（和三种形式）的 Glossoscolex paulistus 血红蛋白（HbGp）的稳定性。观察到 HbGp 寡聚体解离、变性和铁氧化。CD 数据表明，氰合-HbGp 比氧合形式更稳定。氧合和氰合-HbGp 均符合两态模型，在 220-222nm 处的临界脲浓度分别为 5.1±0.2 和 6.1±0.1mol/L。三态模型能够揭示在较低脲浓度（1.0-2.0mol/L）下与部分寡聚体解离相关的微妙的第二个转变。氧合和氰合-HbGp 的中间态与天然态非常相似。对于 met-HbGp，在脲存在下观察到不同的平衡。在 1.95±0.05mol/L 变性剂处观察到明显的转变，与寡聚体解离和亚铁血红素形成有关。在这种情况下，通过三态模型分析揭示了中间态和无规卷曲态之间的巨大相似性。通过两态和三态模型对光谱数据的分析表明，HbGp 脲变性获得的热力学参数具有一致性。
