Section on Physical Biochemistry, Laboratory of Biochemistry and Genetics, National Institute of Diabetes and Digestive and Kidney Diseases, National institutes of Health, U.S. Department of Health and Human Services, Bethesda, Maryland 20892, USA.
J Phys Chem B. 2013 Apr 4;117(13):3554-9. doi: 10.1021/jp4006923. Epub 2013 Mar 25.
An assay for the determination of the equilibrium constant for heteroassociation of α-chymotrypsin and soybean trypsin inhibitor via fluorescence depolarization is described. Results obtained at neutral pH in saline buffer were consistent with prior determinations via sedimentation equilibrium and static light scattering. The dependence of the association equilibrium constant upon the concentrations of urea and trimethylamine-N-oxide (TMAO) added individually and in mixtures was determined at several temperatures. It was found that subdenaturing concentrations of urea decrease the extent of heteroassociation and that added TMAO increases the extent of heteroassociation. The effects of both cosolutes in mixtures upon the equilibrium heteroassociation of α-chymotrypsin and soybean trypsin inhibitor appear to be additive. A thermodynamic analysis of the combined results is presented.
描述了一种通过荧光偏振测定α-糜蛋白酶和大豆胰蛋白酶抑制剂异源缔合平衡常数的分析方法。在中性 pH 值的盐缓冲液中得到的结果与先前通过沉降平衡和静态光散射得到的结果一致。在几个温度下，确定了单独和混合加入尿素和三甲胺 N-氧化物 (TMAO) 时，缔合平衡常数对浓度的依赖性。结果发现，亚变性浓度的尿素降低了异源缔合的程度，而加入的 TMAO 增加了异源缔合的程度。两种共溶剂在混合物中对α-糜蛋白酶和大豆胰蛋白酶抑制剂平衡异源缔合的影响似乎是可加的。给出了对综合结果的热力学分析。
