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How the structure of the large subunit controls function in an oxygen-tolerant [NiFe]-hydrogenase

DOI: 10.1042/BJ20131520 DOI Help
PMID: 24428762 PMID Help

Authors: Lisa Bowman (University of Dundee) , Lindsey Flanagan (University of York) , Paul Fyfe (University of Dundee) , Alison Parkin (University of York) , William Hunter (University of Dundee) , Frank Sargent (University of Dundee)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Biochemical Journal

State: Published (Approved)
Published: January 2014

Open Access Open Access

Abstract: Salmonella enterica is an opportunistic pathogen that produces a [NiFe]-hydrogenase under aerobic conditions. In the present study, genetic engineering approaches were used to facilitate isolation of this enzyme, termed Hyd-5. The crystal structure was determined to a resolution of 3.2 Å and the hydro-genase was observed to comprise associated large and small subunits. The structure indicated that His229 from the large subunit was close to the proximal [4Fe–3S] cluster in the small subunit. In addition, His229 was observed to lie close to a buried glutamic acid (Glu73), which is conserved in oxygen-tolerant hydrogenases. His229 and Glu73 of the Hyd-5 large subunit were found to be important in both hydrogen oxidation activity and the oxygen-tolerance mechanism. Substitution of His229 or Glu73 with alanine led to a loss in the ability of Hyd-5 to oxidize hydrogen in air. Furthermore, the H229A variant was found to have lost the overpotential requirement for activity that is always observed with oxygen-tolerant [NiFe]-hydrogenases. It is possible that His229 has a role in stabilizing the super-oxidized form of the proximal cluster in the presence of oxygen, and it is proposed that Glu73could play a supporting role in fine-tuning the chemistry of His229 to enable this function.

Journal Keywords: Catalysis; Crystallography; X-Ray; Genetic; Glutamic; Histidine; Hydrogen; Hydrogenase; Oxygen; Protein; Salmonella enterica

Subject Areas: Biology and Bio-materials

Instruments: I03-Macromolecular Crystallography

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