Filippova I Y, Lysogorskaya E N, Anisimova V V, Suvorov L I, Oksenoit E S, Stepanov V M
Department of Chemistry, Moscow State University, Russia.
Anal Biochem. 1996 Feb 15;234(2):113-8. doi: 10.1006/abio.1996.0062.
Via a combination of chemical and enzymatic synthesis, new hexapeptide substrates convenient for use in activity assessment of several aspartyl proteinases--porcine pepsin, human pepsin, gastricsin, and cathepsin D--were prepared. These peptide derivatives, o-aminobenzoyl-Ala-Ala-Phe-Phe-Ala-Ala-p-nitroanilide and N-(o-aminobenzoyl-Ala-Ala-Phe-Phe-Ala-Ala)-N'-2,4-dinitrophenyl ethylenediamine, contain a fluorescent o-aminobenzoyl moiety as well as p-nitroaniline or N-2,4-dinitrophenyl ethylenediamine--the groups that cause fluorescence quenching. Aspartyl proteinases hydrolyze the Phe-Phe peptide bond in the substrates, which diminishes quenching due to separation of the fluorescent and quenching moieties and leads to an increase in the fluorescence intensity of o-aminobenzoyl residue. Abz-Ala-Ala-Phe-Phe-Ala-Ala-Ded, being fairly well hydrolyzed by HIV proteinase, might be used for assay of this enzyme.
通过化学合成与酶促合成相结合的方法，制备了几种便于用于多种天冬氨酸蛋白酶（猪胃蛋白酶、人胃蛋白酶、胃泌素和组织蛋白酶D）活性评估的新型六肽底物。这些肽衍生物，邻氨基苯甲酰基 - 丙氨酸 - 丙氨酸 - 苯丙氨酸 - 苯丙氨酸 - 丙氨酸 - 丙氨酸 - 对硝基苯胺和N -（邻氨基苯甲酰基 - 丙氨酸 - 丙氨酸 - 苯丙氨酸 - 苯丙氨酸 - 丙氨酸 - 丙氨酸）- N'-2,4 - 二硝基苯基乙二胺，含有一个荧光邻氨基苯甲酰基部分以及对硝基苯胺或N - 2,4 - 二硝基苯基乙二胺——这些基团会导致荧光猝灭。天冬氨酸蛋白酶水解底物中的苯丙氨酸 - 苯丙氨酸肽键，这由于荧光部分和猝灭部分的分离而减少了猝灭，并导致邻氨基苯甲酰基残基的荧光强度增加。Abz - Ala - Ala - Phe - Phe - Ala - Ala - Ded能被HIV蛋白酶较好地水解，可用于该酶的测定。
