Infrared spectra of amide groups in α-helical proteins:: Evidence for hydrogen bonding between helices and water
Infrared spectra of amide groups in α-helical proteins:: Evidence for hydrogen bonding between helices and water
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DOI：
10.1021/ja001782z
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发表时间：
2000-10-18
影响因子：
15
通讯作者：
Vanderkooi, JM
中科院分区：
文献类型：
--
作者：
Manas, ES;Getahun, Z;Vanderkooi, JM
酰胺振动模的红外光谱频率对二级结构敏感。在这项工作中，提出了水的可获得性另外影响光谱位置的证据。二聚体α-螺旋卷曲的GCN4-P1‘被C-13标记在埋藏的Leu或暴露的Ala的酰胺基团中。在20℃时，C-13丙氨酰胺的酰胺I‘峰位于1585 cm(-1)，而C-13亮氨酸的位置在1606 cm(-1)。这些位移允许区分溶剂暴露的酰胺基团和埋藏的酰胺基团。降低温度会增加氢键的强度，产生向较低频率的转变。在10~273K的甘油水溶液中，与多肽键模型化合物N-甲基乙酰胺在相同的水溶液中类似，分配给螺旋的溶剂暴露区域的酰胺跃迁经历了最强烈的温度依赖的位移。此外，N-甲基乙酰胺的酰胺带和暴露在溶剂中的蛋白质组分的光谱位移随冷冻溶剂的玻璃化转变温度而变化。相反，分配给与水几乎没有相互作用的α-螺旋链段的酰胺转变经历了最弱的位移。α-螺旋蛋白的酰胺I‘带也显示出随温度不同而移动的子峰，并根据它们的温度依赖性和频率被分配到溶剂暴露或埋藏区域。根据氢键强度的变化讨论了光谱位移，包括样品体积膨胀的贡献，以及涉及溶剂和蛋白质的一系列低频振动模引起的平均氢键角的变化。对同位素标记的多肽的结果最终表明，可以通过酰胺I‘峰的位置和依赖于温度的位移来区分暴露在或不暴露于溶剂的α-螺旋区域。
Infrared spectral frequencies of amide vibrational modes are sensitive to secondary structure. In this work, evidence is presented that accessibility to water additionally affects spectral positions. The dimeric alpha -helical coiled-coil GCN4-P1' was C-13 labeled in the amide carbonyl groups of buried Leu or exposed Ala. At 20 degreesC, the amide I' peak for C-13 Ala amide is at 1585 cm(-1), whereas the position for C-13 Leu is at 1606 cm(-1). These shifts permit the distinction of solvent-exposed and buried amide groups. Lowering temperature increases H-bond strength, producing a shift to lower frequency. In the temperature range from 10 to 273 K in aqueous glycerol, the amide transitions assigned to solvent-exposed regions of the helices undergo the strongest temperature-dependent shifts, similar to that of the peptide bond model compound, N-methylacetamide, in the same aqueous solvent. In addition, spectral shifts of the amide bands for N-methylacetamide and the solvent-exposed component of the proteins follow the glass transition temperature of the cryosolvent. In contrast, the amide transitions assigned to alpha -helical segments that are expected to have little interaction with water undergo the weakest shifts. The amide I' band of the alpha -helical protein parvalbumin also shows subpeaks that shift differently with temperature, and on the basis of their temperature dependence and frequency can be assigned to solvent exposed or buried regions. The spectral shifts are discussed in terms of changes in hydrogen bond strengths, including contributions from volume expansion of the sample, and variations in the average hydrogen bond angle, induced by population of low-frequency librational modes involving the solvent and protein. The results on the isotopically labeled peptides conclusively show that alpha -helical regions that are or are not solvent exposed can be distinguished both by the position of the amide I' peak and by the temperature-dependent shifts.