Toomik R, Edlund M, Ek P, Obrink B, Engström L
Department of Medical and Physiological Chemistry, Uppsala University, Sweden.
Pept Res. 1996 Jan-Feb;9(1):6-11.
Sets of peptides with defined sequences, each on a separate spot, were synthesized simultaneously on continuous cellulose membranes (SPOTs membranes), which were originally designed for epitope studies. The applicability of the membrane-bound peptides as substrates for protein kinases was tested using protein kinase A, protein kinase C and casein kinases I and II as model enzymes. We found that the peptide-membrane complexes can serve as kinase substrates. Our results suggest that membrane-bound peptides offer a new potential for the investigation of substrate specificity of protein kinases. An advantage to this method is that there is no need for substrate identification and separation, which is required with high-volume random peptide libraries. Membrane-bound peptides may even form a basis for kinase assays with peptides lacking multiple basic amino acids, required for separation of the substrates in conventional assays. Problems connected with protein kinase substrate specificity can be investigated in any laboratory using the rapid and inexpensive SPOTs technique, as neither costly apparatus nor special experience in peptide synthesis is necessary.
具有特定序列的肽组，每个肽位于单独的斑点上，在连续纤维素膜（SPOTs膜）上同时合成，该膜最初是为表位研究设计的。使用蛋白激酶A、蛋白激酶C以及酪蛋白激酶I和II作为模型酶，测试了膜结合肽作为蛋白激酶底物的适用性。我们发现肽 - 膜复合物可作为激酶底物。我们的结果表明，膜结合肽为研究蛋白激酶的底物特异性提供了新的潜力。这种方法的一个优点是无需进行底物鉴定和分离，而这对于大容量随机肽文库是必需的。膜结合肽甚至可能为缺乏传统测定中底物分离所需多个碱性氨基酸的肽的激酶测定奠定基础。使用快速且廉价的SPOTs技术，任何实验室都可以研究与蛋白激酶底物特异性相关的问题，因为既不需要昂贵的仪器，也不需要肽合成方面的特殊经验。
