UMR 8182, CNRS, Institut de Chimie Moleculaire & des Matériaux d'Orsay, University Paris-Saclay, F-91405 Orsay, France.
Departament de Química, Universitat Autònoma de Barcelona, 08193 Bellaterra, Spain.
Int J Mol Sci. 2022 Nov 24;23(23):14640. doi: 10.3390/ijms232314640.
The covalent insertion of a cobalt heme into the cavity of an artificial protein named alpha Rep (αRep) leads to an artificial cobalt hemoprotein that is active as a catalyst not only for the photo-induced production of H, but also for the reduction of CO in a neutral aqueous solution. This new artificial metalloenzyme has been purified and characterized by Matrix Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS), circular dichroism, and UltraViolet-Visible spectroscopy. Using theoretical experiments, the structure of this biohybrid and the positioning of the residues near the metal complex were examined, which made it possible to complete the coordination of the cobalt ion by an axial glutamine Gln283 ligand. While the Co(III)-porphyrin catalyst alone showed weak catalytic activity for both reactions, 10 times more H and four times more CO were produced when the Co(III)-porphyrin complex was buried in the hydrophobic cavity of the protein. This study thus provides a solid basis for further improvement of these biohybrids using well-designed modifications of the second and outer coordination sphere by site-directed mutagenesis of the host protein.
钴卟啉通过共价键插入名为αRep（αRep）的人工蛋白空腔中，形成了一种人工钴血红素蛋白，这种蛋白不仅可以作为光诱导 H2 产生的催化剂，还可以在中性水溶液中还原 CO。这种新型人工金属酶已经通过基质辅助激光解吸电离飞行时间质谱（MALDI-TOF MS）、圆二色性和紫外可见光谱进行了纯化和表征。通过理论实验，对该生物杂合体的结构和金属配合物附近残基的定位进行了研究，这使得轴向谷氨酰胺 Gln283 配体能够完成钴离子的配位。虽然 Co(III)-卟啉催化剂单独对这两个反应的催化活性都很弱，但当 Co(III)-卟啉配合物埋藏在蛋白质的疏水性腔中时，H2 的产量增加了 10 倍，CO 的产量增加了 4 倍。因此，本研究为进一步通过对宿主蛋白的定点突变对第二和外部配位球进行精心设计的修饰来改进这些生物杂合体提供了坚实的基础。
