Signature of Mobile Hydrogen Bonding of Lysine Side Chains from Long-Range 15N-13C Scalar J-Couplings and Computation

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 N-15-C-13 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 H-1/C-13/N-15 heteronuclear correlation experiments selective for lysine NH3+ groups, we analyzed two different types of long-range N-15-C-13 J-coupling constants: one between intraresidue N-15 zeta and C-13 gamma nuclei ((3)J(N zeta C gamma)) and the other between N-15 zeta and carbonyl C-13' nuclei across a hydrogen bond ((h3)J(N zeta C')). The experimental (3)J(N zeta C gamma) data confirm the highly mobile nature of the chi(4) torsion angles of lysine side chains seen in the MD simulation. The NH3+ groups of Lys29 and Lys33 exhibit measurable (h3)J(N zeta C') couplings arising from hydrogen bonds with backbone carbonyl groups of Glu16 and Thr14, respectively. When interpreted together with the (3)J(N zeta C gamma)-coupling constants and NMR-relaxation-derived S-2 order parameters of the NH3+ groups, they strongly suggest that hydrogen bonds involving NH3+ groups are of a transient and highly dynamic nature, in remarkably good agreement with the MD simulation results.