Journal
ANGEWANDTE CHEMIE-INTERNATIONAL EDITION
Volume 57, Issue 33, Pages 10605-10609Publisher
WILEY-V C H VERLAG GMBH
DOI: 10.1002/anie.201805144
Keywords
enzyme catalysis; FTIR spectroscopy; hydride species; hydrogenases; nuclear resonance vibrational spectroscopy
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Funding
- NIH [GM-65440]
- Cluster of Excellence Unifying Concepts in Catalysis initiative of the DFG [EXC 314]
- U.S. Department of Energy (DOE) [DE-AC36-08GO28308]
- U.S. Department of Energy, Office of Basic Energy Sciences, Division of Chemical Sciences, Geosciences, and Biosciences
- NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES [R01GM065440] Funding Source: NIH RePORTER
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A combination of nuclear resonance vibrational spectroscopy (NRVS), FTIR spectroscopy, and DFT calculations was used to observe and characterize Fe-H/D bending modes in CrHydA1 [FeFe]-hydrogenase Cys-to-Ser variant C169S. Mutagenesis of cysteine to serine at position169 changes the functional group adjacent to the H-cluster from a -SH to -OH, thus altering the proton transfer pathway. The catalytic activity of C169S is significantly reduced compared to that of native CrHydA1, presumably owing to less efficient proton transfer to the H-cluster. This mutation enabled effective capture of a hydride/deuteride intermediate and facilitated direct detection of the Fe-H/D normal modes. We observed a significant shift to higher frequency in an Fe-H bending mode of the C169S variant, as compared to previous findings with reconstituted native and oxadithiolate (ODT)-substituted CrHydA1. On the basis of DFT calculations, we propose that this shift is caused by the stronger interaction of the -OH group of C169S with the bridgehead -NH- moiety of the active site, as compared to that of the -SH group of C169 in the native enzyme.
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