Koenig S H, Beaulieu C F, Brown R D, Spiller M
IBM T. J. Watson Research Center, Yorktown Heights, New York 10598.
Biophys J. 1990 Mar;57(3):461-9. doi: 10.1016/S0006-3495(90)82562-5.
From analyses of the magnetic field dependence of 1/T1 (nuclear magnetic relaxation dispersion [NMRD] profiles) of water protons in solutions of highly purified calf lens gamma II-crystallin, we find that monomers form oligomers at relatively low concentrations, which increase in size with increasing concentration and decreasing temperature. At approximately 16% by volume and -4 degrees C, the mean oligomeric molecular weight is approximately 120-fold greater than the monomeric value of 20 kD. Below this concentration, there is no indication of any substantive change in conformation of the monomeric subunits. At higher concentrations, the tertiary structure of the monomer appears to reconfigure rather abruptly, but reversibly, as evidenced by the appearance of spectra-like 14N peaks in the NMRD profiles. The magnitudes of these peaks, known to arise from cross-relaxation of water protons through access to amide (NH) moieties of the protein backbone, indicate that the high concentration conformation is not compact, but open and extended in a manner that allows enhanced interaction with solvent. The data are analogous to those found for homogenates of calf and chicken lens (Beaulieu, C. F., J. I. Clark, R. D. Brown III, M. Spiller, and S. H. Koenig. 1988. Magn. Reson. Med. 8:47-57; Beaulieu, C. F., R. D. Brown III, J. I. Clark, M. Spiller, and S. H. Koenig. 1989. Magn. Reson. Med. 10:62-72). This unusually large dependence of oligomeric size and conformation on concentration in the physiological range is suggested as the mechanism by which osmotic equilibrium is maintained, at minimal metabolic expense, in the presence of large gradients of protein concentration in the lens in vivo (cf Vérétout and Tardieu, 1989. Eur. Biophys. J. 17:61-68). Finally, the results of the NMRD data provide a ready explanation of the low temperature phase transition, and "cold-cataract" separation of phases, observed in gamma II-crystallin solutions; we suggest that the phases that separate are the two major conformers detected by NMRD.
通过对高纯度小牛晶状体γII-晶状体蛋白溶液中水质子的1/T1（核磁共振弛豫色散[NMRD]谱）的磁场依赖性分析，我们发现单体在相对较低浓度下形成寡聚体，其大小随浓度增加和温度降低而增大。在约16%（体积）和-4℃时，平均寡聚体分子量比20kD的单体值大约大120倍。低于此浓度时，没有迹象表明单体亚基的构象有任何实质性变化。在较高浓度下，单体的三级结构似乎相当突然但可逆地重新构型，NMRD谱中出现类似光谱的14N峰就证明了这一点。这些峰的大小已知是由于水质子通过与蛋白质主链的酰胺（NH）基团接触而发生交叉弛豫产生的，这表明高浓度构象不是紧凑的，而是开放且伸展的，从而允许与溶剂增强相互作用。这些数据与在小牛和鸡晶状体匀浆中发现的数据类似（Beaulieu, C. F., J. I. Clark, R. D. Brown III, M. Spiller, and S. H. Koenig. 1988. Magn. Reson. Med. 8:47 - 57; Beaulieu, C. F., R. D. Brown III, J. I. Clark, M. Spiller, and S. H. Koenig. 1989. Magn. Reson. Med. 10:62 - 72）。这种寡聚体大小和构象在生理范围内对浓度的异常大的依赖性被认为是在体内晶状体中存在大的蛋白质浓度梯度时，以最小的代谢代价维持渗透平衡的机制（参见Vérétout和Tardieu, 1989. Eur. Biophys. J. 17:61 - 68）。最后，NMRD数据的结果为在γII-晶状体蛋白溶液中观察到的低温相变和“冷白内障”相分离提供了现成的解释；我们认为分离出的相是NMRD检测到的两种主要构象体。
