Department of Chemistry, University of California, Davis, CA 95616, USA.
J Inorg Biochem. 2010 Oct;104(10):1063-70. doi: 10.1016/j.jinorgbio.2010.06.003. Epub 2010 Jul 1.
Proton 2D NMR was used to confirm in solution a highly conserved portion of the molecular structure upon substrate loss for the heme oxygenase from the pathogenic bacterium Corynebacterium diphtheriae, HmuO. The chemical shifts for the conserved portion of the structure are assessed as references for the dipolar shifts needed to determine the orientation of the paramagnetic susceptibility tensor, chi, in paramagnetic substrate complexes of HmuO. It is shown that the chemical shifts for the structurally conserved portion of substrate-free HmuO serve as excellent references for residues with only small to moderate sized dipolar shifts in the cyanide-inhibited substrate complex of HmuO, yielding an orientation of chi that is essentially the same as conventionally obtained from large dipolar shifts based on empirical estimates of the diamagnetic reference. The implications of these diamagnetic chemical shifts for characterizing the hydrogen bonding in the physiologically relevant, resting-state, high-spin aquo complex are discussed. The pattern of labile proton exchange in the distal H-bond network of substrate-free HmuO allowed comparison of changes in dynamic stability of tertiary contacts in the substrate-free and substrate-bound HmuO and with the same complexes of human heme oxygenase.
利用质子二维 NMR 技术，在 Corynebacterium diphtheriae 致病细菌血红素加氧酶失去底物的情况下，对其结构中高度保守的部分进行了溶液中的确认。结构中保守部分的化学位移被评估为确定 HmuO 顺磁底物复合物中顺磁磁化率张量 chi 的取向所需的偶极位移的参考值。结果表明，无底物 HmuO 结构保守部分的化学位移可作为 HmuO 氰化物抑制底物复合物中偶极位移较小或中等的残基的极好参考，从而产生与传统方法相同的 chi 取向，后者基于顺磁参考的经验估计的大偶极位移。这些抗磁化学位移对表征生理相关的、静止状态的高自旋水合复合物中的氢键的意义进行了讨论。无底物 HmuO 中远端氢键网络中不稳定质子交换的模式允许比较无底物和底物结合的 HmuO 以及与人血红素加氧酶相同的复合物中三级接触的动态稳定性变化。
