Department of Chemistry, Lancaster University, Lancaster, LA1 4YB, UK.
WestCHEM, Department of Pure & Applied Chemistry, University of Strathclyde, Glasgow, G1 1XL, UK.
Solid State Nucl Magn Reson. 2018 Aug;92:7-11. doi: 10.1016/j.ssnmr.2018.03.003. Epub 2018 Mar 14.
Nuclear magnetic resonance (NMR) is a powerful tool for investigating atomic-scale structure in heterogeneous or composite materials where long-range order is absent. In this work solid-state H and H-detected NMR experiments were performed with fast magic angle spinning (ν = 75 kHz) and at high magnetic fields (B = 20 T) and used to gain structural insight into a heterogeneous biocatalyst consisting of an enzyme, human carbonic anhydrase II (hCA II), covalently immobilized on epoxy-functionalized silica. Two-dimensional H-H NOESY-type correlation experiments were able to provide information on H environments in silica, epoxy-silica and the immobilized enzyme. Two distinct signals originating from water protons were observed: water associated with the surface of the silica and the water associated with the immobilized enzyme. Additional two-dimensional H-H double-single quantum (DQ-SQ) correlation experiments suggested that the immobilized enzyme is not in close contact with the silica surface. Most significantly, comparison of two-dimensional H-N spectra of the immobilized enzyme and the solution-state enzyme confirmed that the structural integrity of the protein is well preserved upon covalent immobilization.
核磁共振（NMR）是一种强大的工具，可用于研究缺乏长程有序的非均相或复合材料中的原子尺度结构。在这项工作中，使用快速魔角旋转（ν=75 kHz）和高磁场（B=20 T）进行了固态 H 和 H 检测 NMR 实验，以深入了解由酶（人碳酸酐酶 II（hCA II））共价固定在环氧功能化硅上的非均相生物催化剂的结构。二维 H-H NOESY 型相关实验能够提供有关二氧化硅、环氧硅和固定化酶中 H 环境的信息。观察到两个源自水质子的不同信号：与二氧化硅表面相关的水和与固定化酶相关的水。另外，二维 H-H 双单量子（DQ-SQ）相关实验表明，固定化酶与二氧化硅表面没有密切接触。最重要的是，比较固定化酶和溶液态酶的二维 H-N 谱证实，蛋白质的结构完整性在共价固定化后得到很好的保留。
