Mast cell tryptases: examination of unusual characteristics by multiple sequence alignment and molecular modeling.
Mast cell tryptases: examination of unusual characteristics by multiple sequence alignment and molecular modeling.
复制标题
肥大细胞类胰蛋白酶：通过多重序列比对和分子建模检查异常特征。
DOI：
10.1002/pro.5560010309
复制
发表时间：
1992
期刊：
影响因子：
--
通讯作者：
Barton,GJ
中科院分区：
文献类型：
--
作者：
Johnson,DA;Barton,GJ
胰蛋白酶是在肥大细胞颗粒中发现的胰蛋白酶样丝氨酸蛋白酶。虽然它们显示出与胰蛋白酶40%的序列同一性，并且仅含有20或21个额外的残基，但胰蛋白酶显示出几个不寻常的特征。与胰蛋白酶不同，胰蛋白酶仅在少数蛋白质中进行有限的切割，并且不受天然胰蛋白酶抑制剂的抑制，它们形成四聚体，结合肝素，并且它们对合成底物的活性随着盐浓度的增加而逐渐受到抑制高于0.2 M。通过与其他丝氨酸蛋白酶进行比较，鉴定了七种胰蛋白酶的独特序列特征。然后通过基于牛胰蛋白酶晶体结构的分子建模预测胰蛋白酶的三维结构。该模型显示两个大的插入位于活性位点裂缝的两侧，这表明了对蛋白质底物上的胰蛋白酶活性有限以及缺乏天然抑制剂抑制的解释。一组保守的Trp残基和一个独特的富含脯氨酸的区域形成两个表面疏水补丁，这可能是四聚体形成和/或盐浓度增加抑制的原因。虽然它们不含共有的肝素结合序列，但胰蛋白酶比胰蛋白酶多10-13个His残基，并且这些残基位于模型的表面上。此外，Arg和Lys残基的聚集也可能有助于肝素结合。推定的Asn连接糖基化位点位于模型活性位点的相对侧。该模型为胰蛋白酶的一些不寻常特征提供了结构解释，并为未来的实验提供了合理的基础，例如定点突变。
Tryptases are trypsin‐like serine proteinases found in the granules of mast cells. Although they show 40% sequence identity with trypsin and contain only 20 or 21 additional residues, tryptases display several unusual features. Unlike trypsin, the tryptases only make limited cleavages in a few proteins and are not inhibited by natural trypsin inhibitors, they form tetramers, bind heparin, and their activity on synthetic substrates is progressively inhibited as the concentration of salt increases above 0.2 M.Unique sequence features of seven tryptases were identified by comparison to other serine proteinases. The three‐dimensional structures of the tryptases were then predicted by molecular modeling based on the crystal structure of bovine trypsin. The models show two large insertions to lie on either side of the active‐site cleft, suggesting an explanation for the limited activity of tryptases on protein substrates and the lack of inhibition by natural inhibitors. A group of conserved Trp residues and a unique proline‐rich region make two surface hydrophobic patches that may account for the formation of tetramers and/or inhibition with increasing salt. Although they contain no consensus heparin‐binding sequence, the tryptases have 10–13 more His residues than trypsin, and these are positioned on the surface of the model. In addition, clustering of Arg and Lys residues may also contribute to heparin binding. Putative Asn‐linked glycosylation sites are found on the opposite side of the model from the active site. The model provides structural explanations for some to the unusual characteristics of the tryptases and a rational basis for future experiments, such as site‐directed mutagenesis.