Lee Y C, Jackson P L, Jablonsky M J, Muccio D D, Pfister R R, Haddox J L, Sommers C I, Anantharamaiah G M, Chaddha M
Department of Chemistry, University of Alabama at Birmingham, 901 S. 14th Street, Birmingham, AL 35294, USA.
Biopolymers. 2001 May;58(6):548-61. doi: 10.1002/1097-0282(200105)58:6<548::AID-BIP1030>3.0.CO;2-B.
Alkaline hydrolysis of corneal proteins in the alkali-injured eye releases N-acetyl-proline-glycine-proline (Ac-Pro-Gly-Pro-OH) among other peptides. It has been shown that this tripeptide is a neutrophil chemoattractant. Existing data suggest that the release of this peptide is the catalytic event for early neutrophil invasion of the cornea leading to corneal ulcers. In order to design inhibitors of this tripeptide chemoattractant that would block neutrophil invasion and diminish corneal ulcers, we studied the solution properties of this tripeptide by NMR spectroscopy and compared this peptide to Ac-Pro-Gly-OH (a weaker chemoattractant), and to Ac-Pro-OH (inactive). The NMR data were consistent with Ac-Pro-Gly-Pro-OH existing in solution as a mixture of four isomers with different cis and trans conformations about the two X-proline amide bonds. The isomer with two trans conformations (trans-trans) was the most dominant (41%) in aqueous solution. This was followed by the isomers with mixed cis and trans conformations (trans-cis, 26% and cis-trans, 20%). The isomer with two cis conformations (cis-cis) was the least favored (13%). The populations of these isomers were investigated in DMSO and they were similar to those reported in aqueous solutions except that the ordering of the trans-cis and cis-trans isomers were reversed. NMR NH temperature coefficients and nuclear Overhauser effect (NOE) measurements as well as CD spectroscopy were used to demonstrate that the four isomers exist primarily in an extended conformation with little hydrogen bonding. The available (NOE) information was used with molecular dynamics calculations to construct a dominant solution conformation for each isomer of the tripeptide. This information will serve as a model for the design of peptide and nonpeptide inhibitors of the chemoattractant.
在碱烧伤的眼睛中，角膜蛋白的碱性水解会释放出N - 乙酰 - 脯氨酸 - 甘氨酸 - 脯氨酸（Ac - Pro - Gly - Pro - OH）以及其他肽段。研究表明，这种三肽是一种中性粒细胞趋化因子。现有数据表明，该肽的释放是中性粒细胞早期侵入角膜导致角膜溃疡的催化事件。为了设计能够阻断中性粒细胞侵入并减轻角膜溃疡的这种三肽趋化因子抑制剂，我们通过核磁共振光谱研究了该三肽的溶液性质，并将其与Ac - Pro - Gly - OH（一种较弱的趋化因子）和Ac - Pro - OH（无活性）进行了比较。核磁共振数据表明，Ac - Pro - Gly - Pro - OH在溶液中以四种异构体的混合物形式存在，这四种异构体围绕两个X - 脯氨酸酰胺键具有不同的顺式和反式构象。具有两个反式构象（反 - 反）的异构体在水溶液中占主导地位（41%）。其次是具有顺式和反式混合构象的异构体（反 - 顺，26%和顺 - 反，20%）。具有两个顺式构象（顺 - 顺）的异构体最不受青睐（13%）。我们在二甲基亚砜中研究了这些异构体的比例，结果与在水溶液中报道的相似，只是反 - 顺和顺 - 反异构体的顺序颠倒了。核磁共振NH温度系数、核Overhauser效应（NOE）测量以及圆二色光谱被用于证明这四种异构体主要以伸展构象存在，氢键作用很少。利用现有的（NOE）信息结合分子动力学计算，构建了该三肽每种异构体的主要溶液构象。这些信息将作为设计趋化因子的肽类和非肽类抑制剂的模型。
