Bell J E, Castellino F J, Trayer I P, Hill R L
J Biol Chem. 1975 Oct 10;250(19):7579-85.
Reaction of alpha-lactalbumin at pH 7 in aqueous solution with either 2-hydroxy-5-nitrobenzylbromide or N-bromosuccinimide yields derivatives in which only 2 of the 4 tryptophan residues are modified. All 4 residues of tryptophan are modified under the similar conditions in 8 M urea. Structural analysis of the modified derivatives revealed that tryptophans 26 and 118 are the sole reactive residues and that tryptophan 118 reacts more rapidly than tryptophan 26. The fluorescence of alpha-lactalbumin modified to varying extents with N-bromosuccinimide indicates that tryptophan 118 is exposed to solvent whereas tryptophan 26 is in a more hydrophobic environment. The chemical reactivities and fluorescence properties of tryptophans 26 and 118 are consistent with the proposed conformations of alpha-lactalbumin based on its similarity with egg white lysozyme. The kinetic properties of both derivatives of alpha-lactalbumin containing up to 2 modified residues indicate that each derivative has decreased affinity for the galactosyltransferase but that at saturating concentrations, Km and Vmax for lactose synthesis are unchanged from those of native alpha-lactalbumin.
α-乳白蛋白在pH 7的水溶液中与2-羟基-5-硝基苄基溴或N-溴代琥珀酰亚胺反应，生成的衍生物中4个色氨酸残基只有2个被修饰。在8M尿素的类似条件下，所有4个色氨酸残基都被修饰。对修饰衍生物的结构分析表明，色氨酸26和118是唯一的反应性残基，且色氨酸118比色氨酸26反应更快。用N-溴代琥珀酰亚胺不同程度修饰的α-乳白蛋白的荧光表明，色氨酸118暴露于溶剂中，而色氨酸26处于更疏水的环境中。色氨酸26和118的化学反应性和荧光特性与基于其与蛋清溶菌酶相似性所提出的α-乳白蛋白构象一致。两种含有多达2个修饰残基的α-乳白蛋白衍生物的动力学特性表明，每种衍生物对半乳糖基转移酶的亲和力都降低了，但在饱和浓度下，乳糖合成的Km和Vmax与天然α-乳白蛋白的相同。
