Sun T X, Akhtar N J, Liang J J
Center for Ophthalmic Research, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts 02115, USA.
J Biol Chem. 1999 Nov 26;274(48):34067-71. doi: 10.1074/jbc.274.48.34067.
Lens alpha-crystallin is a 600-800-kDa heterogeneous oligomer protein consisting of two subunits, alphaA and alphaB. The homogeneous oligomers (alphaA- and alphaB-crystallins) have been prepared by recombinant DNA technology and shown to differ in the following biophysical/biochemical properties: hydrophobicity, chaperone-like activity, subunit exchange rate, and thermal stability. In this study, we studied their thermodynamic stability by unfolding in guanidine hydrochloride. The unfolding was probed by three spectroscopic parameters: absorbance at 235 nm, Trp fluorescence intensity at 320 nm, and far-UV circular dichroism at 223 nm. Global analysis indicated that a three-state model better describes the unfolding behavior than a two-state model, an indication that there are stable intermediates for both alphaA- and alphaB-crystallins. In terms of standard free energy (DeltaG(NU)(H(2)(O))), alphaA-crystallin is slightly more stable than alphaB-crystallin. The significance of the intermediates may be related to the functioning of alpha-crystallins as chaperone-like molecules.
晶状体α-晶状体蛋白是一种由αA和αB两个亚基组成的600 - 800 kDa异源寡聚体蛋白。通过重组DNA技术制备了均一寡聚体（αA-和αB-晶状体蛋白），并显示它们在以下生物物理/生化特性方面存在差异：疏水性、伴侣样活性、亚基交换率和热稳定性。在本研究中，我们通过在盐酸胍中展开来研究它们的热力学稳定性。通过三个光谱参数来探测展开过程：235 nm处的吸光度、320 nm处的色氨酸荧光强度以及223 nm处的远紫外圆二色性。全局分析表明，三态模型比两态模型更能描述展开行为，这表明αA-和αB-晶状体蛋白都存在稳定的中间体。就标准自由能（ΔG(NU)(H₂O)）而言，αA-晶状体蛋白比αB-晶状体蛋白略稳定。中间体的意义可能与α-晶状体蛋白作为伴侣样分子的功能有关。
