Antenna domain mobility and enzymatic reaction of L-rhamnulose-1-phosphate aldolase
Antenna domain mobility and enzymatic reaction of L-rhamnulose-1-phosphate aldolase
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DOI：
10.1021/bi7012799
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发表时间：
2008-01-15
期刊：
影响因子：
2.9
通讯作者：
Schulz, Georg E.
中科院分区：
文献类型：
--
作者：
Grueninger, Dirk;Schulz, Georg E.
来自大肠杆菌的 L-鼠李糖-1-磷酸醛缩酶参与 L-鼠李糖（一种普遍存在的脱氧己糖）的降解途径。它是一种罕见的 C-4 对称型同四聚体，其 N 末端结构域像天线一样从主体上突出。酶的迁移率分析提出了各向异性热天线运动可能支持催化作用的假设（Kroemer 等人，Biochemistry 42, 10560, 2003）。我们通过生成四个单突变体和一个二硫键来检验这一假设，这些突变体和二硫键旨在降低天线域的移动性而不干扰链折叠或活性中心。突变体的催化速率显示活性降低，这与预期的天线固定密切相关。在这些突变体中，K15W 被结晶、结构阐明，并用作其他突变体建模的指南。该结构证实了设计，因为突变引入了与相邻子单元的紧密非极性接触，固定了天线但不影响主链。通过比较描述域移动性的各向异性 B 因子来确认固定。事实证明，野生型天线域的明显各向异性移动性在 K15W 中已变为各向同性，与设计一致。我们建议，与 K15W 一样，其他突变也遵循该设计，验证天线移动性和活动之间的相关性。这种相关性表明域移动性促进了反应。
The enzyme L-rhamnulose-1-phosphate aldolase from Escherichia coli participates in the degradation pathway Of L-rhamnose, a ubiquitous deoxy-hexose. It is a homotetramer of the rare C-4-symmetric type with N-terminal domains protruding like antennas from the main body. A mobility analysis of the enzyme gave rise to the hypothesis that an anisotropic thermal antenna motion may support the catalysis (Kroemer et A, Biochemistry 42, 10560, 2003). We checked this hypothesis by generating four single mutants and one disulfide bridge that were designed to reduce the mobility of the antenna domain without disturbing the chain-fold or the active center. The catalytic rates of the mutants revealed activity reductions that correlated well with the expected antenna fixation. Among these mutants, K15W was crystallized, structurally elucidated, and used as a guide for modeling the others. The structure confirmed the design because the mutation introduced a tight nonpolar contact to a neighboring Subunit that fixed the antenna but did not affect the main chain. The fixation was confirmed by a comparison of the anisotropic B-factors describing the mobility of the domains. It turned out that the distinctly anisotropic mobility of the wild-type antenna domain has become isotropic in K15W, in agreement with the design. We suggest that, like K15W, the other mutations also followed the design, validating the correlation between antenna mobility and activity. This correlation suggests that the domain mobility facilitates the reaction.