Department of Chemical Engineering, Stanford University, Stanford, CA, 94305, USA.
Department of Bioengineering, Stanford University, Stanford, CA, 94305, USA.
Angew Chem Int Ed Engl. 2017 Jan 19;56(4):1012-1016. doi: 10.1002/anie.201610260. Epub 2016 Dec 15.
Hydrogenases, ferredoxins, and ferredoxin-NADP reductases (FNR) are redox proteins that mediate electron metabolism in vivo, and are also potential components for biological H production technologies. A high-throughput H production assay device (H PAD) is presented that enables simultaneous evaluation of 96 individual H production reactions to identify components that improve performance. Using a CCD camera and image analysis software, H PAD senses the chemo-optical response of Pd/WO thin films to the H produced. H PAD-enabled discovery of hydrogenase and FNR mutants that enhance biological H production is reported. From a library of 10 080 randomly mutated Clostridium pasteurianum [FeFe] hydrogenases, we found a mutant with nearly 3-fold higher H production specific activity. From a library of 400 semi-randomly mutated Oryza sativa FNR, the top hit enabled a 60 % increase in NADPH-driven H production rates. H PAD can also facilitate elucidation of fundamental biochemical mechanisms within these systems.
氢化酶、铁氧还蛋白和铁氧还蛋白-NADP 还原酶（FNR）是介导体内电子代谢的氧化还原蛋白，也是生物制氢技术的潜在组成部分。本文介绍了一种高通量制氢分析装置（H PAD），该装置可同时评估 96 个独立的制氢反应，以识别提高性能的组件。该装置利用 CCD 相机和图像分析软件来感知 Pd/WO 薄膜对产生的 H 的化学-光学响应。利用 H PAD 发现了能够提高生物制氢的氢化酶和 FNR 突变体。从 10080 个随机突变的巴氏梭菌[FeFe]氢化酶文库中，我们发现了一个具有近 3 倍更高的比产氢活性的突变体。从一个 400 个半随机突变的水稻 FNR 文库中，最佳突变体使 NADPH 驱动的产氢速率提高了 60%。H PAD 还可以促进这些系统中基本生化机制的阐明。
