Abdel Malak C, Filippova I Y, Lysogorskaya E N, Anisimova V V, Lavrenova G I, Stepanov V M
Chemistry Department, Moscow State University, USSR.
Int J Pept Protein Res. 1992 May;39(5):443-9. doi: 10.1111/j.1399-3011.1992.tb01448.x.
Pepsin successfully catalyzed the synthesis of several peptide derivatives from N-protected di- or tripeptides and amino acid or peptide esters or p-nitroanilides in dimethylformamide-water solutions at pH 4.6. An optimal substrates:pepsin ratio depended on the structure of starting peptides, especially their fit to the substrate binding sites of the enzyme. For hexapeptide Z-Ala-Ala-Phe-Leu-Ala-Ala-OCH3 formation, an equilibrium yield was attained at 1:3.10(5) enzyme-substrates ratio that indicated high efficiency of pepsin in synthesis reactions. In the course of the equilibrium peptide synthesis, pepsin gradually disappeared from the liquid phase due to its entrapment within a gel, formed by the hexapeptide product, while retaining its activity. The inclusion into the precipitate was not specific for pepsin, so far as inert proteins, lysozyme, ribonuclease A and carbonic anhydrase, when added to the reaction mixture, became also co-precipitated with the hexapeptide formed. It appears that co-precipitation of pepsin, an important factor limiting the enzyme efficiency, might be operative as well for other proteinases used to catalyze peptide synthesis.
在pH 4.6的二甲基甲酰胺-水溶液中，胃蛋白酶成功地催化了由N-保护的二肽或三肽与氨基酸、肽酯或对硝基苯胺合成几种肽衍生物。最佳底物与胃蛋白酶的比例取决于起始肽的结构，尤其是它们与酶的底物结合位点的契合度。对于六肽Z-Ala-Ala-Phe-Leu-Ala-Ala-OCH3的形成，在酶与底物比例为1:3.10(5)时达到平衡产率，这表明胃蛋白酶在合成反应中具有高效率。在平衡肽合成过程中，胃蛋白酶由于被困在由六肽产物形成的凝胶中而逐渐从液相中消失，但其活性得以保留。沉淀中的包裹对胃蛋白酶并非特异性的，因为当向反应混合物中加入惰性蛋白质、溶菌酶、核糖核酸酶A和碳酸酐酶时，它们也会与形成的六肽一起共沉淀。看来，胃蛋白酶的共沉淀是限制酶效率的一个重要因素，对于用于催化肽合成的其他蛋白酶可能也起作用。
