Department of Biochemistry and Molecular Biology, Sealy Center for Structural Biology and Molecular Biophysics, University of Texas Medical Branch, Galveston, Texas 77555, USA.
J Am Chem Soc. 2011 Jun 22;133(24):9192-5. doi: 10.1021/ja202219n. Epub 2011 May 27.
Amino acid side chains involved in hydrogen bonds and electrostatic interactions are crucial for protein function. However, detailed investigations of such side chains in solution are rare. Here, through the combination of long-range (15)N-(13)C scalar J-coupling measurements and an atomic-detail molecular dynamics (MD) simulation, direct insight into the structural dynamic behavior of lysine side chains in human ubiquitin has been gained. On the basis of (1)H/(13)C/(15)N heteronuclear correlation experiments selective for lysine NH(3)(+) groups, we analyzed two different types of long-range (15)N-(13)C J-coupling constants: one between intraresidue (15)Nζ and (13)Cγ nuclei ((3)J(NζCγ)) and the other between (15)Nζ and carbonyl (13)C' nuclei across a hydrogen bond ((h3)J(NζC')). The experimental (3)J(NζCγ) data confirm the highly mobile nature of the χ(4) torsion angles of lysine side chains seen in the MD simulation. The NH(3)(+) groups of Lys29 and Lys33 exhibit measurable (h3)J(NζC') couplings arising from hydrogen bonds with backbone carbonyl groups of Glu16 and Thr14, respectively. When interpreted together with the (3)J(NζCγ)-coupling constants and NMR-relaxation-derived S(2) order parameters of the NH(3)(+) groups, they strongly suggest that hydrogen bonds involving NH(3)(+) groups are of a transient and highly dynamic nature, in remarkably good agreement with the MD simulation results.
涉及氢键和静电相互作用的氨基酸侧链对于蛋白质功能至关重要。然而，在溶液中对这些侧链进行详细研究的情况很少。在这里，通过长程（15）N-（13）C 标量 J 耦合测量和原子细节分子动力学（MD）模拟的结合，直接了解了人泛素中赖氨酸侧链的结构动态行为。基于对赖氨酸 NH（3）（+）基团具有选择性的（1）H/（13）C/（15）N 异核相关实验，我们分析了两种不同类型的长程（15）N-（13）C J 耦合常数：一种是在残基内（15）Nζ和（13）Cγ核之间（（3）J（NζCγ）），另一种是在氢键（（h3）J（NζC'））中穿过（15）Nζ和羰基（13）C'核之间。实验（3）J（NζCγ）数据证实了 MD 模拟中赖氨酸侧链的 χ（4）扭转角具有高度可移动的性质。Lys29 和 Lys33 的 NH（3）（+）基团表现出可测量的（h3）J（NζC'）耦合，分别来自与 Glu16 和 Thr14 的骨架羰基的氢键。当与（3）J（NζCγ）-耦合常数和 NH（3）（+）基团的 NMR 弛豫衍生 S（2）顺序参数一起解释时，它们强烈表明涉及 NH（3）（+）基团的氢键具有瞬态和高度动态的性质，与 MD 模拟结果非常吻合。
