Effect of immobilization site on the orientation and activity of surface-tethered enzymes
Effect of immobilization site on the orientation and activity of surface-tethered enzymes
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DOI：
10.1039/c7cp06063g
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发表时间：
2018-01-14
影响因子：
3.3
通讯作者：
Chen, Zhan
中科院分区：
文献类型：
--
作者：
Li, Yaoxin;Ogorzalek, Tadeusz L.;Chen, Zhan
将肽和蛋白质拴系到非生物表面具有产生具有有用性质的生物分子官能化表面的潜力。常用的固定方法缺乏对生物分子共价或物理结合到表面的取向的控制，导致次优材料。在这里，我们使用了一种工程β-半乳糖苷酶，它可以通过独特的表面可接近的半胱氨酸残基以明确的方向化学固定在表面上。使用和频产生（SFG）振动光谱和粗粒分子动力学（MD）模拟的组合研究进行，以确定酶的固定化位点和非生物表面化学对酶的表面取向，表面覆盖度和催化活性的影响。研究了通过在位置227和308处引入的半胱氨酸固定化的两种β-半乳糖苷酶变体。在这两种情况下，当非生物表面变得更加亲水时，酶表面覆盖率降低，但活性增加。分子动力学模拟表明，这是由于固定化酶和更亲水的表面之间的相互作用减弱。这些研究提供了更好的理解如何酶表面相互作用可以优化，以最大限度地提高表面拴系酶的催化活性。
Tethering peptides and proteins to abiotic surfaces has the potential to create biomolecule-functionalized surfaces with useful properties. Commonly used methods of immobilization lack control over the orientation in which biological molecules are covalently or physically bound to the surface, leading to sub-optimal materials. Here we use an engineered beta-galactosidase that can be chemically immobilized on a surface with a well-defined orientation through unique surface-accessible cysteine residues. A combined study using sum frequency generation (SFG) vibrational spectroscopy and coarse grained molecular dynamics (MD) simulations was performed to determine the effects of enzyme immobilization site and abiotic surface chemistry on enzyme surface orientation, surface coverage, and catalytic activity. Two beta-galactosidase variants that were immobilized through cysteine introduced at positions 227 and 308 were studied. In both cases, when the abiotic surface was made more hydrophilic, the enzyme surface coverage decreased, but the activity increased. MD simulations indicated that this is due to the weakened interactions between the immobilized enzyme and the more hydrophilic surface. These studies provide improved understanding of how enzyme-surface interactions can be optimized to maximize the catalytic activity of surface tethered enzymes.