Characterization of the key step for light-driven hydrogen evolution in green algae.
Under anaerobic conditions, several species of green algae perform a light-dependent hydrogen production catalyzed by a special group of [FeFe] hydrogenases termed HydA. Although highly interesting for biotechnological applications, the direct connection between photosynthetic electron transport and hydrogenase activity is still a matter of speculation. By establishing an in vitro reconstitution system, we demonstrate that the photosynthetic ferredoxin (PetF) is essential for efficient electron transfer between photosystem I and HydA1. To investigate the electrostatic interaction process and electron transfer between PetF and HydA1, we performed site-directed mutagenesis. Kinetic analyses with several site-directed mutagenesis variants of HydA1 and PetF enabled us to localize the respective contact sites. These experiments in combination with in silico docking analyses indicate that electrostatic interactions between the conserved HydA1 residue Lys(396) and the C terminus of PetF as well as between the PetF residue Glu(122) and the N-terminal amino group of HydA1 play a major role in complex formation and electron transfer. Mapping of relevant HydA1 and PetF residues constitutes an important basis for manipulating the physiological photosynthetic electron flow in favor of light-driven H(2) production.
Lehrstuhl Biochemie der Pflanzen, AG Photobiotechnologie, Ruhr-Universität Bochum, Universitätsstrasse 150, 44801 Bochum, Germany.
Martin
M
Winkler
Institut für Biochemie und Biotechnologie der Pflanzen, Universität Münster, Hindenburgplatz 55, 49143 Münster, Germany.
Sebastian
S
Kuhlgert
Institut für Biochemie und Biotechnologie der Pflanzen, Universität Münster, Hindenburgplatz 55, 49143 Münster, Germany.
Michael
M
Hippler
Lehrstuhl Biochemie der Pflanzen, AG Photobiotechnologie, Ruhr-Universität Bochum, Universitätsstrasse 150, 44801 Bochum, Germany. Electronic address: thomas.happe@rub.de.
Thomas
T
Happe
D020418:Algal Proteins / Q000235:genetics / Q000378:metabolism*; D000693:Anaerobiosis / Q000502:physiology; D000818:Animals; D016825:Chlamydomonas reinhardtii / Q000201:enzymology* / Q000235:genetics; D004579:Electron Transport / Q000502:physiology; D005288:Ferredoxins / Q000235:genetics / Q000378:metabolism; D006859:Hydrogen / Q000378:metabolism*; D006864:Hydrogenase / Q000235:genetics / Q000378:metabolism*; D008027:Light; D010449:Peptide Mapping; D010788:Photosynthesis / Q000502:physiology*; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism*
D020418:Algal Proteins; D005288:Ferredoxins; D015800:Protozoan Proteins; D006859:Hydrogen; D006864:Hydrogenase