Oxygen Pathways and Allostery in Monomeric Sarcosine Oxidase via Single-Sweep Free-Energy Reconstruction.
Oxygen Pathways and Allostery in Monomeric Sarcosine Oxidase via Single-Sweep Free-Energy Reconstruction.
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DOI：
10.1021/ct500088z
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发表时间：
2014-07-08
影响因子：
5.5
通讯作者：
Abrams, Cameron F.
中科院分区：
文献类型：
--
作者：
Bucci, Anthony;Abrams, Cameron F.
单体肌氨酸氧化酶(MSOX)是一种黄素蛋白D-氨基酸氧化酶，其肌氨酸和氧的活化位点分别位于黄素环的Re和Si面上。O2向催化内部的传输途径还不是很清楚，仅从MSOX的晶体结构很难确定。一种称为单扫描法的复合自由能方法被用来绘制和热力学表征氧位和从蛋白质表面通向具有催化活性的Lys265的路线。结果是MSOX内部的路径和自由能网络，说明氧可以从四个单独的溶剂入口访问还原黄素表面的两个自由能极小值。在硅表面上没有观察到这样的最小值。该酶的三种主要状态的途径在几何上是相似的：(1)具有闭合黄素裂隙的载脂蛋白，(2)具有开放黄素裂隙的载脂蛋白，以及(3)与抑制剂结合的闭合黄素裂隙。有趣的是，当底物模拟抑制剂2-呋喃甲酸结合在肌氨酸位点时，这些运输途径上的自由能显示出明显更深的最小值，即使在远离该位点的位置上也是如此。这表明O2从溶剂到活性中心的转运动力学依赖于底物的变构调节。
Monomeric sarcosine oxidase (MSOX) is a flavoprotein D-amino acid oxidase with reported sarcosine and oxygen activation sites on the re and si faces of the flavin ring, respectively. O2 transport routes to the catalytic interior are not well understood and are difficult to ascertain solely from MSOX crystal structures. A composite free-energy method known as single-sweep is used to map and thermodynamically characterize oxygen sites and routes leading to the catalytically active Lys265 from the protein surface. The result is a network of pathways and free energies within MSOX illustrating that oxygen can access two free-energy minima on the re face of the reduced flavin from four separate solvent portals. No such minimum is observed on the si face. The pathways are geometrically similar for three major states of the enzyme: (1) apo with a closed flavin cleft, (2) apo with an open flavin cleft, and (3) inhibitor-bound with a closed flavin cleft. Interestingly, free energies along these transport pathways display significantly deeper minima when the substrate-mimicking inhibitor 2-furoic acid is bound at the sarcosine site, even at locations far from this site. This suggests a substrate-dependent allosteric modulation of the kinetics of O2 transport from the solvent to the active site.
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