CHARACTERIZATION OF A PARTLY FOLDED PROTEIN BY NMR METHODS - STUDIES ON THE MOLTEN GLOBULE STATE OF GUINEA-PIG ALPHA-LACTALBUMIN
CHARACTERIZATION OF A PARTLY FOLDED PROTEIN BY NMR METHODS - STUDIES ON THE MOLTEN GLOBULE STATE OF GUINEA-PIG ALPHA-LACTALBUMIN
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DOI：
10.1021/bi00427a002
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发表时间：
1989-01-10
期刊：
影响因子：
2.9
通讯作者：
HANLEY, C
中科院分区：
文献类型：
--
作者：
BAUM, J;DOBSON, CM;HANLEY, C
NMR光谱学已被用于研究蛋白质的部分折叠状态的结构，α-淀粉酶的熔融球或A-状态。乳白蛋白。该物种的1H NMR谱与天然和完全未折叠状态的1H NMR谱有很大不同，反映了中间水平的顺序。光谱中的分辨率受到许多共振的广泛重叠和实质线宽的限制。因此，已经开发了利用天然蛋白质的良好分辨光谱来间接探测A状态的方法。已经发现A状态的许多共振从它们在展开状态的谱中的位置基本上偏移，并且已经在两个状态相互转换的条件下通过与原生状态的磁化转移来识别。在A-状态中最强烈扰动的残基被发现是形成天然结构的疏水核心的那些残基中的B。发现许多酰胺在A-状态下被高度保护免于溶剂交换。这些已经通过pH跳跃实验鉴定，该实验在天然蛋白质的光谱中标记它们。他们被发现主要发生在天然结构中的螺旋段。这些结果使一个模型的A-状态被提出，其中存在显着的构象自由，但保留了特定的元素的本地样结构。
NMR spectroscopy has been used to investigate the structure of a partially folded state of a protein, the molten globule or A-state of .alpha.-lactalbumin. The 1H NMR spectrum of this species differs substantially from those of both the native and fully unfolded states, reflecting the intermediate level of order. The resolution in the spectrum is limited by the widespread overlap and substantial line widths of many of the resonances. Methods have therefore been developed that exploit the well-resolved spectrum of the native protein to probe indirectly the A-state. A number of resonances of the A-state have been found to be substantially shifted from their positions in the spectrum of the unfolded state and have been identified through magnetization transfer with the native state, under conditions where the two states are interconverting. The most strongly perturbed residues in the A-state were found to b eamong those that form a hydrophobic core to the native structure. A number of amides were found to be highly protected from solvent exchange in the A-state. These have been identified through pH-jump experiments, which label them in the spectrum of the native protein. They were found to occur mainly in segments that are helical in the native structure. These results enable a model of the A-state to be proposed in which significant conformational freedom exists but where specific elements of native-like structure are preserved.