Alcohol-induced protein folding transitions in platelet factor 4: the O-state.
Alcohol-induced protein folding transitions in platelet factor 4: the O-state.
复制标题
酒精诱导血小板因子 4 中的蛋白质折叠转变：O 状态。
DOI：
10.1021/bi00084a038
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发表时间：
1993
期刊：
影响因子：
2.9
通讯作者：
Mayo,KH
中科院分区：
文献类型：
--
作者：
Yang,Y;Mayo,KH
摘要：血小板因子4 (PF4)（7800道尔顿）是一种反平行/3-sheet， a//3类蛋白，其三级结构通过两个二硫键的存在而稳定。用2-丙醇或类似的低分子量脂肪醇滴定PF4可诱导可逆的蛋白质折叠转变，在600 mhz 'H NMR时间尺度上观察到这种转变是缓慢交换的。对Y60、H35、H23的环状质子和K50在天然状态和醇诱导状态（o态）下的aH的分辨共振进行线拟合，可以推导出折叠平衡常数和交换动力学。从9.8 M - 2-丙醇到3.3 M - 2-丙醇，折叠汇率在5到100 s_1之间变化。对0 M 2-丙醇进行简单的线性外推，得到o态到n态的交换速率约为500 s_1，即毫秒时间尺度。在相对较高的2-丙醇浓度下（> 9.5 M），其中o态占主导地位（> 90%），核磁共振光谱显示出更“展开”的结构，而CD数据表明保留了相当多的二级结构。然而，将2-丙醇从3.3 M增加到9.8 M， cd衍生的分数组成显著变化，总/3结构减少约20%，a-螺旋组成增加约25%。醇跳实验在原生PF4的核磁共振光谱中发现了o态长寿命NHs，表明大约15种原生和o态长寿命NHs的折叠转变可逆性和保守性。大多数这些国民保健服务被分配到残基在反平行/3-片结构。在这15种NHs中，H/D汇率虽然在o态有所降低，但与随机线圈H/D交换相比，通常仍然长期存在。总的来说，pf4 o态是一个稳定的中间体，具有明显更高的反平行/3-片结构波动和更稳定的c端a-螺旋。
Revised Manuscript Received May 10, 1993 abstract: Platelet factor 4 (PF4)(7800 daltons) is an anti-parallel/3-sheet, a//3 class protein whose tertiary structure is stabilized by the presence of two disulfide bonds. Titration of PF4 with 2-propanol or similar low molecular weight, aliphatic alcohols induces reversible proteinfolding transitions which are observed to be in slow exchange on the 600-MHz'H NMR time scale. Line fitting of resolved resonances assigned to ring protons of Y60, H35, H23, and aH of K50 in native and alcohol-induced states (O-states) allows derivation of folding equilibrium constants and exchange kinetics. Folding exchange rates vary between 5 and 100 s_1 on going from 9.8 to 3.3 M 2-propanol. Simple linear extrapolation to 0 M 2-propanol yields an O-state to N-state exchange rate of about 500 s_1, ie, millisecond time scale. At relativelyhigh 2-propanol concentration (> 9.5 M), where the O-state is predominant (> 90%), NMR spectra suggest a more “unfolded” structure, while CD data indicate the preservation of considerable secondary structure. Increasing 2-propanol from 3.3 to 9.8 M, however, shifts the CD-derivedfractional compositions significantly, with overall/3-structure decreasing by about 20% and a-helix compositionincreasing by about 25%. Alcohol-jump experiments, whichidentify O-state long-lived NHs in the NMR spectrum of native PF4, indicate folding transition reversibility and conservation of about 15 long-lived NHs in native and O-states. Most of these NHs are assigned to residues in anti-parallel/3-sheet structure. Of these 15 NHs, H/D exchange rates, althoughvariably reduced in the O-state, are generally still long-lived compared with random coil H/D exchange. Overall, thePF4 O-state is a stable intermediate with an apparently more highly fluctuating anti-parallel/3-sheet structure and a more stabilized C-terminal a-helix.