Wolfson A J, Shrimpton C N, Lew R A, Smith A I
Peptide Biology Laboratory, Baker Medical Research Institute, Prahran, Victoria, Australia.
Biochem Biophys Res Commun. 1996 Dec 4;229(1):341-8. doi: 10.1006/bbrc.1996.1803.
The activity of endopeptidase EC 3.4.24.15 (thimet oligopeptidase, EP 24.15), as measured by cleavage of a quenched fluorescent substrate, 7-methoxycoumarin-4-acetyl-Pro-Leu-Gly-Pro-D-Lys (2,4-dinitrophenyl), was increased 2-3 fold by the addition of 1 mM Mn2+ or of 10 mM Ca2+. The inhibitory capability of a specific EP. 24.15 inhibitor, N-[1-(R,S)-carboxy-3-phenylpropyl]-Ala-Ala-Tyr-p-aminobenzoate, was also increased at similar concentrations of these metal ions. However, the hydrolysis of naturally-occurring peptides, thought to be the physiological substrates for EP 24.15, was not affected by either Mn2+ or Ca2+. These results suggest that the binding of synthetic analogs to the enzyme may differ significantly from the binding, and thus hydrolysis, of natural peptide substrates and caution against drawing conclusions about substrate interactions with the active site from data obtained with modified peptide ligands.
通过切割淬灭荧光底物7-甲氧基香豆素-4-乙酰基-脯氨酸-亮氨酸-甘氨酸-脯氨酸-D-赖氨酸（2,4-二硝基苯基）来测定的内肽酶EC 3.4.24.15（硫醚内肽酶，EP 24.15）的活性，在添加1 mM Mn2+或10 mM Ca2+后增加了2至3倍。特异性EP 24.15抑制剂N-[1-(R,S)-羧基-3-苯基丙基]-丙氨酸-丙氨酸-酪氨酸-对氨基苯甲酸酯的抑制能力在这些金属离子的相似浓度下也有所增加。然而，被认为是EP 24.15生理底物的天然存在的肽的水解不受Mn2+或Ca2+的影响。这些结果表明，合成类似物与酶的结合可能与天然肽底物的结合以及因此的水解有显著差异，并提醒不要从用修饰的肽配体获得的数据得出关于底物与活性位点相互作用的结论。
