500-MHz1H NMR studies of insulin: Complete assignment of histidine resonances
500-MHz1H NMR studies of insulin: Complete assignment of histidine resonances
复制标题
胰岛素的 500-MHz1H NMR 研究：组氨酸共振的完整分配
DOI：
10.1007/bf01025598
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发表时间：
1983
期刊：
影响因子：
--
通讯作者：
K. Williamson
中科院分区：
文献类型：
--
作者：
Dalia Cheshovsky;L. Neuringer;K. Williamson
胰岛素单体的两个组氨酸在将胰岛素组织成胰岛素六聚体中起着至关重要的作用。B10组氨酸与锌结合形成双锌胰岛素六聚体，B5和B10组氨酸均参与形成四锌胰岛素六聚体。这两种组氨酸都可接近二聚体形式胰岛素的溶剂，在pH 2-3下存在的主要物质。在目前的工作中，我们报告的第一个500兆赫1H NMR研究胰岛素。在该频率下，来自每个等效单体的两个组氨酸的所有四个质子共振都被分辨。共振归属于B5 His和B10 His的C（2）-和C（4）-咪唑质子，采用Carr-Purcell脉冲序列检测单线态并观察近似T2弛豫时间。在高达60°C的温度下，在乙酸盐缓冲液和不同浓度的尿素中检查pH 2.9的无锌牛胰岛素。B5 His在二聚体的分子I和分子II中的环境必须是相同的，B10 His也是如此，因为总共只看到四个尖锐的共振。我们对两个C（2）质子的分配与最近对人（B5 Ala）胰岛素的研究结果一致。
The two histidines of the insulin monomer play a vital role in the organization of insulin into insulin hexamers. The B10 histidines bind to zinc to form two-zinc insulin hexamer, and both the B5 and B10 histidines are implicated in the formation of four-zinc insulin hexamer. These two histidines are both accessible to solvent in the dimeric form of insulin, the predominant species present at pH 2–3. In the present work we report the first 500-MHz1H NMR studies of insulin. At this frequency all four proton resonances from the two histidines of each equivalent monomer are resolved. The resonances are assigned to the C(2)- and C(4)-imidazole protons of B5 His and B10 His employing Carr-Purcell pulse sequences to detect singlets and to observe approximateT2 relaxation times. Zinc-free bovine insulin at pH 2.9 was examined at temperatures up to 60°C in acetate buffer and in urea of varying concentrations. The environments of B5 His in molecule I and molecule II of the dimer must be the same, with the same being true for B10 His, since a total of only four sharp resonances are seen. Our assignments for the two C(2) protons are consistent with those determined from recent studies of human (B5 Ala) insulin.