Eftink M R, Ghiron C A
Biochemistry. 1977 Dec 13;16(25):5546-51. doi: 10.1021/bi00644a024.
The acrylamide quenching reaction is shown to be very discriminating in sensing the exposure of fluorescing tryptophanyl residues in globular proteins. The quenching rate constants for some proteins, such as aldolase and human serum albumin, are reported to be independent of the solvent viscosity, indicating that the reaction is limited by penetration of the quencher through the protein matrix. Temperature-dependent studies are performed to determine the activation energy and entropy for the penetration of acrylamide into these proteins. The tryptophanyl residues in aldolase are shown to be shielded by a large activation energy barrier, while the single residue in human serum albumin is shielded by a large activation entropy barrier. These parameters characterize the nature of the protein matrix enveloping the fluorophors.
丙烯酰胺猝灭反应在检测球状蛋白质中荧光色氨酸残基的暴露情况时表现出很强的选择性。据报道，一些蛋白质（如醛缩酶和人血清白蛋白）的猝灭速率常数与溶剂粘度无关，这表明该反应受猝灭剂穿透蛋白质基质的限制。进行了温度依赖性研究，以确定丙烯酰胺渗透到这些蛋白质中的活化能和熵。结果表明，醛缩酶中的色氨酸残基受到较大活化能垒的屏蔽，而人血清白蛋白中的单个残基则受到较大活化熵垒的屏蔽。这些参数表征了包裹荧光团的蛋白质基质的性质。
