Goehlert Virginia A, Krupinska Ewa, Regan Lynne, Stone Martin J
Department of Chemistry, Indiana University, Bloomington, Indiana 47405-0001, USA.
Protein Sci. 2004 Dec;13(12):3322-30. doi: 10.1110/ps.04926604. Epub 2004 Nov 10.
"Host-guest" studies of the B1 domain from Streptococcal protein G have been used previously to establish a thermodynamic scale for the beta-sheet-forming propensities of the 20 common amino acids. To investigate the contribution of side chain conformational entropy to the relative stabilities of B1 domain mutants, we have determined the dynamics of side chain methyl groups in 10 of the 20 mutants used in a previous study. Deuterium relaxation rates were measured using two-dimensional NMR techniques for 13CH2D groups. Analysis of the relaxation data using the Lipari-Szabo model-free formalism showed that mutations introduced at the guest position caused small but statistically significant changes in the methyl group dynamics. In addition, there was a low level of covariation of the Lipari-Szabo order parameters among the 10 mutants. The variations in conformational free energy estimated from the order parameters were comparable in magnitude to the variations in global stability of the 10 mutants but did not correlate with the global stability of the domain or with the structural properties of the guest amino acids. The data support the view that conformational entropy in the folded state is one of many factors that can influence the folding thermodynamics of proteins.
先前已利用来自链球菌蛋白G的B1结构域的“主客体”研究，为20种常见氨基酸的β-折叠形成倾向建立了一个热力学标度。为了研究侧链构象熵对B1结构域突变体相对稳定性的贡献，我们测定了先前一项研究中使用的20个突变体中10个突变体侧链甲基的动力学。使用二维核磁共振技术测量了13CH2D基团的氘弛豫率。使用Lipari-Szabo无模型形式对弛豫数据进行分析表明，在客体位置引入的突变导致甲基动力学发生了微小但具有统计学意义的变化。此外，10个突变体之间Lipari-Szabo序参数的协变水平较低。根据序参数估计的构象自由能变化幅度与10个突变体的全局稳定性变化相当，但与结构域的全局稳定性或客体氨基酸的结构特性无关。这些数据支持了这样一种观点，即折叠态的构象熵是影响蛋白质折叠热力学的众多因素之一。
