NMR study suggests a major role for Arg111 in maintaining the structure and dynamical properties of type II human cellular retinoic acid binding protein.
NMR study suggests a major role for Arg111 in maintaining the structure and dynamical properties of type II human cellular retinoic acid binding protein.
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NMR 研究表明 Arg111 在维持 II 型人细胞视黄酸结合蛋白的结构和动力学特性方面发挥着重要作用。
DOI：
10.1021/bi981021x
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发表时间：
1998
期刊：
影响因子：
--
通讯作者：
Yan,H
中科院分区：
文献类型：
--
作者：
Wang,L;Yan,H
通过核磁共振光谱法确定了Arg 111被甲硫氨酸（R111 M）取代的II型人细胞视黄酸结合蛋白（CRABPII）定点突变体的溶液结构。用多核多维NMR对apo-R111 M的1H和15 N共振峰进行了顺序归属。从NMR数据中得到的2302个距离约束和77个π-二面角约束计算溶液结构。28个代表结构的精细构象体的系综的均方根偏差对于主链原子和所有重原子分别为0.54 ± 0.26和0.98 ± 0.23 π。apo-R111 M的溶液结构与野生型apo-CRABPII相似。然而，两种蛋白质之间存在显着的构象差异，主要集中在三个片段（Leu 19 − Ala 36，Glu 73 − Cys 81和Leu 99 − Pro 105），这些片段聚集在距离点突变超过17 bp的配体入口周围。在apo-R111 M中，所有三个片段都向配体入口的中心移动，使得apo-R111 M中配体结合口袋的开口比野生型apo-CRABPII中的小得多。此外，apo-R111 M的配体结合口袋，特别是配体入口，比apo-CRABPII的柔性小得多。令人惊讶的是，apo-R111 M在结构和动力学性质上都更类似于holo-CRABPII而不是apo-CRABPII。突变引起的构象和动力学变化与RA结合引起的变化相似，但突变引起的变化幅度小于RA结合引起的变化。结果表明，Arg 111在决定CRABPII的结构和动力学性质方面起着关键作用。
The solution structure of a site-directed mutant of type-II human cellular retinoic acid binding protein (CRABPII) with Arg111 replaced by methionine (R111M) has been determined by NMR spectroscopy. The sequential assignments of the1H and15N resonances of apo-R111M were established by multinuclear multidimensional NMR. The solution structure was calculated from 2302 distance restraints and 77 ϕ dihedral restraints derived from the NMR data. The root-mean-square deviation of the ensemble of 28 refined conformers that represent the structure from the mean coordinate set derived from them was 0.54 ± 0.26 and 0.98 ± 0.23 Å for the backbone atoms and all heavy atoms, respectively. The solution structure of apo-R111M is similar to that of wild-type apo-CRABPII. However, there are significant conformational differences between the two proteins, localized mainly to three segments (Leu19−Ala36, Glu73−Cys81, and Leu99−Pro105) clustered around the ligand entrance more than 17 Å away from the point mutation. In apo-R111M, all the three segments move toward the center of the ligand entrance so that the opening of the ligand-binding pocket in apo-R111M is much smaller than that in wild-type apo-CRABPII. Furthermore, the ligand-binding pocket of apo-R111M, especially the ligand entrance, is much less flexible than that of apo-CRABPII. Surprisingly, apo-R111M is more similar to holo-CRABPII than to apo-CRABPII in both structure and dynamical properties. The conformational and dynamical changes caused by the mutation are similar to those induced by binding of RA, although the magnitudes of the changes caused by the mutation are smaller than those induced by binding of RA. The results suggest that Arg111 plays a critical role in determining the structure and dynamical properties of CRABPII.