Kaplan A P, Bartlett P A
Department of Chemistry, University of California, Berkeley 94720.
Biochemistry. 1991 Aug 20;30(33):8165-70. doi: 10.1021/bi00247a011.
Comparative studies among a series of tripeptide phosphonate inhibitors of the zinc peptidase carboxypeptidase A indicate that incorporation of the phosphonic acid analogue of valine at the P1 position results in significantly higher affinity than the glycine, alanine, or phenylalanine analogues. When applied to the tripeptide analogue Cbz-Phe-ValP-(O)Phe [ZFVP(O)F], determination of the inhibition constant Ki was complicated by the very slow rate of dissociation. The rate of exchange of [3H]ZFVP(O)F with enzyme-bound [14C]ZFVP(O)F was followed for periods of 3-4 months to measure dissociation rate constants in the range of (1.7-4.4) x 10(-9) s-1, corresponding to half-lives of 5-13 years. Although the on- and off-rate constants differ for different carboxypeptidase isozymes, their ratios, corresponding to the inhibition constants Ki, are consistently in the range of 10-27 fM. Both the inhibition constants and the dissociation rate constants appear to be the lowest values yet determined for an enzyme-small inhibitor interaction.
一系列锌肽酶羧肽酶A的三肽膦酸酯抑制剂的比较研究表明，在P1位置引入缬氨酸的膦酸类似物会导致其亲和力显著高于甘氨酸、丙氨酸或苯丙氨酸类似物。当应用于三肽类似物Cbz-Phe-ValP-(O)Phe [ZFVP(O)F]时，抑制常数Ki的测定因解离速率非常缓慢而变得复杂。跟踪[3H]ZFVP(O)F与酶结合的[14C]ZFVP(O)F的交换速率长达3 - 4个月，以测量解离速率常数，其范围为(1.7 - 4.4)×10(-9)s-1，对应半衰期为5 - 13年。尽管不同羧肽酶同工酶的结合和解离速率常数不同，但其比值（对应于抑制常数Ki）始终在10 - 27 fM范围内。抑制常数和解离速率常数似乎都是目前所测定的酶 - 小抑制剂相互作用的最低值。
