Martin M, Vallee B L, Riordan J F
Anal Biochem. 1987 Mar;161(2):341-7. doi: 10.1016/0003-2697(87)90460-x.
Dansylated tight-binding inhibitors are effective fluorophoric probes for detecting conformational changes of enzyme active sites. In this study they have been employed to examine the effect of anions on the conformation of angiotensin-converting enzyme. The efficiency of radiationless energy transfer between enzyme tryptophan residues and an active site-bound dansyl inhibitor has been shown to be enhanced by the addition of chloride. Half-maximal fluorescence enhancement occurs at about 2 mM chloride and is the same for both N-(1-carboxyl-5-dansylamino-pentyl)-glycyl-L-phenylalanine [Ki,app = 50 nM (pH 7.5, 300 mM NaCl)] and N-(1-carboxyl-5-dansylamino-pentyl)-glycyl-L-lysine (Ki,app = 5.7 nM). Other activating anions also evoke similar increases in enzyme-inhibitor energy transfer. Fluorescence changes are not due to binding additional inhibitor molecules but rather to an anion-induced change in protein conformation.
丹磺酰化紧密结合抑制剂是用于检测酶活性位点构象变化的有效荧光探针。在本研究中，它们已被用于研究阴离子对血管紧张素转换酶构象的影响。已表明，通过添加氯离子，酶色氨酸残基与活性位点结合的丹磺酰抑制剂之间的无辐射能量转移效率会提高。在约2 mM氯离子浓度时出现最大荧光增强的一半，并且对于N-(1-羧基-5-丹磺酰氨基戊基)-甘氨酰-L-苯丙氨酸[Ki,app = 50 nM（pH 7.5，300 mM NaCl）]和N-(1-羧基-5-丹磺酰氨基戊基)-甘氨酰-L-赖氨酸（Ki,app = 5.7 nM）而言是相同的。其他活化阴离子也会引起酶-抑制剂能量转移的类似增加。荧光变化不是由于结合了额外的抑制剂分子，而是由于阴离子诱导的蛋白质构象变化。
