Study of nucleic acid structure by novel NMR methods

当寡核苷酸在磁场中排列时，13C-1H双重态的下场TROSY分量由于残余偶极13C-1H偶极耦合(RDC)和残余13C化学位移各向异性(RCSA)而改变其共振频率，这两个二级张量之和称为伪CSA。实验测得的取向和各向同性样品中13C-TROSY分量的共振频率的差异称为剩余伪CSA(RPCSA)，它可以直接作为结构计算的约束。由于RPCSA的测量涉及到窄TROSY 13C双线分量的检测，因此它适用于具有比RDC测量更大的旋转相关时间的系统。该方法被证明用于24-NT茎环片段或核糖体螺旋35的螺旋区的结构精细化，该螺旋区在13C和15N中均匀富含，RPCSA值在5℃和25℃测量，在加入RPCSA限制后，结构准确性得到了显著的交叉验证改善。

Upon alignment of oligonucleotides in a magnetic field, the downfield, TROSY-component of the 13C-1H doublet changes its resonance frequency as a result of residual dipolar 13C-1H dipolar coupling (RDC) and residual 13C chemical shift anisotropy (RCSA), and the sum of these two second rank tensors is referred to as the pseudo-CSA. The experimentally measured difference in the resonance frequency of the 13C TROSY component in the aligned and isotropic samples is referred to as residual pseudo-CSA (RPCSA), and it can be used directly as a restraint during structure calculation. Because measurement of the RPCSA involves detection of the narrow TROSY 13C doublet component, it is applicable to systems with larger rotational correlation times than RDC measurement. The method is demonstrated for structure refinement of the helical region of a 24-nt stem-loop segment or ribosomal helix 35, uniformly enriched in 13C and 15N, with RPCSA values measured at 5 and 25 C. Substantial cross-validated improvements in structural accuracy are obtained upon incorporation of RPCSA restraints.