Department of Chemistry and Biochemistry, The Ohio State University, 100 W 18th Avenue, Columbus, Ohio 43210, United States.
Biochemistry. 2023 Sep 5;62(17):2622-2631. doi: 10.1021/acs.biochem.3c00249. Epub 2023 Aug 14.
Nickel-substituted rubredoxin (NiRd) from has previously been shown to act as both a structural and functional mimic of the [NiFe] hydrogenase. However, improvements both in turnover frequency and overpotential are needed to rival the native [NiFe] hydrogenase enzymes. Characterization of a library of NiRd mutants with variations in the secondary coordination sphere suggested that protein dynamics played a substantial role in modulating activity. In this work, rubredoxin scaffolds were selected from diverse organisms to study the effects of distal sequence variation on catalytic activity. It was found that though electrochemical catalytic activity was only slightly impacted across the series, the Rd sequence from a psychrophilic organism exhibited substantially higher levels of solution-phase hydrogen production. Additionally, Eyring analyses suggest that catalytic activation properties relate to the growth temperature of the parent organism, implying that the general correlation between the parent organism environment and catalytic activity often seen in naturally occurring enzymes may also be observed in artificial enzymes. Selecting protein scaffolds from hosts that inhabit diverse environments, particularly low-temperature environments, represents an alternative approach for engineering artificial metalloenzymes.
镍取代豆血红蛋白（NiRd）来自先前已被证明可以作为 [NiFe] 氢化酶的结构和功能模拟物。然而，需要提高周转率和过电势才能与天然 [NiFe] 氢化酶相媲美。对具有不同二级配位球的 NiRd 突变体文库的表征表明，蛋白质动力学在调节活性方面起着重要作用。在这项工作中，从不同的生物体中选择豆血红蛋白支架来研究远端序列变异对催化活性的影响。结果发现，尽管电化学催化活性在整个系列中仅略有影响，但来自嗜冷生物的 Rd 序列表现出更高水平的溶液相产氢。此外，埃林分析表明，催化激活特性与母体生物的生长温度有关，这意味着在天然存在的酶中经常观察到的母体生物环境与催化活性之间的一般相关性也可能在人工酶中观察到。从栖息在不同环境（特别是低温环境）中的宿主中选择蛋白质支架，是工程化人工金属酶的另一种方法。
