Dedman J R, Kaetzel M A, Chan H C, Nelson D J, Jamieson G A
Department of Physiology and Biophysics, University of Cincinnati, College of Medicine, Ohio 45267.
J Biol Chem. 1993 Nov 5;268(31):23025-30.
The interaction of short amino acid sequences is the basis of molecular recognition and biological regulation in many cellular systems. Libraries of random peptides provide an approach to identify peptides that can be used to modulate, in a targeted fashion, the function of specific gene products. We have used a library of random peptides designed and constructed in the M13 bacteriophage to select calcium-dependent calmodulin binding-peptides. Twenty-eight independent sequences were obtained; all contained a tryptophan within the fifteen-amino acid insert. In 11 sequences, the tryptophan was located in the first possible variable position of the inserted sequence and was followed by a proline. The tryptophan-proline combination was also present in six additional isolates but at various other positions within the peptide insert. Synthetic peptides, representative of the calmodulin binding sequences, bound to calmodulin in a calcium-dependent fashion, competed with known calmodulin inhibitors and, when introduced via a patch pipette, inhibited calcium-activated chloride conductance of the colonic epithelial cell line, T84. This report demonstrates the utility of identifying modifiers of biological function and should prove to be a valuable approach in understanding the cellular role of proteins of unknown function.
短氨基酸序列之间的相互作用是许多细胞系统中分子识别和生物调节的基础。随机肽文库提供了一种方法，可用于鉴定能够以靶向方式调节特定基因产物功能的肽。我们利用在M13噬菌体中设计和构建的随机肽文库来筛选钙依赖性钙调蛋白结合肽。获得了28个独立序列；所有序列在15个氨基酸插入片段中都含有一个色氨酸。在11个序列中，色氨酸位于插入序列的第一个可能的可变位置，并紧跟一个脯氨酸。色氨酸-脯氨酸组合也存在于另外6个分离株中，但位于肽插入片段内的其他不同位置。代表钙调蛋白结合序列的合成肽以钙依赖性方式与钙调蛋白结合，与已知的钙调蛋白抑制剂竞争，并且当通过膜片钳微管引入时，抑制结肠上皮细胞系T84的钙激活氯电导。本报告证明了鉴定生物功能调节剂的实用性，并且应该证明是理解未知功能蛋白质细胞作用的一种有价值的方法。
