title: Crystal structure and functional characterization of photosystem II-associated carbonic anhydrase CAH3 in Chlamydomonas reinhardtii.
abstract: In oxygenic photosynthesis, light energy is stored in the form of chemical energy by converting CO2 and water into carbohydrates. The light-driven oxidation of water that provides the electrons and protons for the subsequent CO2 fixation takes place in photosystem II (PSII). Recent studies show that in higher plants, HCO3 (-) increases PSII activity by acting as a mobile acceptor of the protons produced by PSII. In the green alga Chlamydomonas reinhardtii, a luminal carbonic anhydrase, CrCAH3, was suggested to improve proton removal from PSII, possibly by rapid reformation of HCO3 (-) from CO2. In this study, we investigated the interplay between PSII and CrCAH3 by membrane inlet mass spectrometry and x-ray crystallography. Membrane inlet mass spectrometry measurements showed that CrCAH3 was most active at the slightly acidic pH values prevalent in the thylakoid lumen under illumination. Two crystal structures of CrCAH3 in complex with either acetazolamide or phosphate ions were determined at 2.6- and 2.7-Å resolution, respectively. CrCAH3 is a dimer at pH 4.1 that is stabilized by swapping of the N-terminal arms, a feature not previously observed in α-type carbonic anhydrases. The structure contains a disulfide bond, and redox titration of CrCAH3 function with dithiothreitol suggested a possible redox regulation of the enzyme. The stimulating effect of CrCAH3 and CO2/HCO3 (-) on PSII activity was demonstrated by comparing the flash-induced oxygen evolution pattern of wild-type and CrCAH3-less PSII preparations. We showed that CrCAH3 has unique structural features that allow this enzyme to maximize PSII activity at low pH and CO2 concentration.
authors.affiliation: Department of Forest Genetics and Plant Physiology (R.B) and Department of Plant Physiology (T.S., G.S.), Umeå Plant Science Centre, and Department of Chemistry, Chemistry Biology Centre (D.S., T.H., C.G., J.M., A.E.S.-E.), Umeå University, SE-90187 Umea, Sweden.
authors.forename: Reyes
authors.identifier: http://orcid.org/0000-0002-0364-1589
authors.initials: R
authors.lastname: Benlloch
authors.affiliation: Department of Forest Genetics and Plant Physiology (R.B) and Department of Plant Physiology (T.S., G.S.), Umeå Plant Science Centre, and Department of Chemistry, Chemistry Biology Centre (D.S., T.H., C.G., J.M., A.E.S.-E.), Umeå University, SE-90187 Umea, Sweden.
authors.forename: Dmitriy
authors.initials: D
authors.lastname: Shevela
authors.affiliation: Department of Forest Genetics and Plant Physiology (R.B) and Department of Plant Physiology (T.S., G.S.), Umeå Plant Science Centre, and Department of Chemistry, Chemistry Biology Centre (D.S., T.H., C.G., J.M., A.E.S.-E.), Umeå University, SE-90187 Umea, Sweden.
authors.forename: Tobias
authors.initials: T
authors.lastname: Hainzl
authors.affiliation: Department of Forest Genetics and Plant Physiology (R.B) and Department of Plant Physiology (T.S., G.S.), Umeå Plant Science Centre, and Department of Chemistry, Chemistry Biology Centre (D.S., T.H., C.G., J.M., A.E.S.-E.), Umeå University, SE-90187 Umea, Sweden.
authors.forename: Christin
authors.initials: C
authors.lastname: Grundström
authors.affiliation: Department of Forest Genetics and Plant Physiology (R.B) and Department of Plant Physiology (T.S., G.S.), Umeå Plant Science Centre, and Department of Chemistry, Chemistry Biology Centre (D.S., T.H., C.G., J.M., A.E.S.-E.), Umeå University, SE-90187 Umea, Sweden.
authors.forename: Tatyana
authors.initials: T
authors.lastname: Shutova
authors.affiliation: Department of Forest Genetics and Plant Physiology (R.B) and Department of Plant Physiology (T.S., G.S.), Umeå Plant Science Centre, and Department of Chemistry, Chemistry Biology Centre (D.S., T.H., C.G., J.M., A.E.S.-E.), Umeå University, SE-90187 Umea, Sweden.
authors.forename: Johannes
authors.initials: J
authors.lastname: Messinger
authors.affiliation: Department of Forest Genetics and Plant Physiology (R.B) and Department of Plant Physiology (T.S., G.S.), Umeå Plant Science Centre, and Department of Chemistry, Chemistry Biology Centre (D.S., T.H., C.G., J.M., A.E.S.-E.), Umeå University, SE-90187 Umea, Sweden goran.samuelsson@umu.se elisabeth.sauer-eriksson@umu.se.
authors.forename: Göran
authors.initials: G
authors.lastname: Samuelsson
authors.affiliation: Department of Forest Genetics and Plant Physiology (R.B) and Department of Plant Physiology (T.S., G.S.), Umeå Plant Science Centre, and Department of Chemistry, Chemistry Biology Centre (D.S., T.H., C.G., J.M., A.E.S.-E.), Umeå University, SE-90187 Umea, Sweden goran.samuelsson@umu.se elisabeth.sauer-eriksson@umu.se.
authors.forename: A Elisabeth
authors.initials: AE
authors.lastname: Sauer-Eriksson
mesh_terms: D002257:Carbonic Anhydrase Inhibitors / Q000494:pharmacology; D002256:Carbonic Anhydrases / Q000737:chemistry* / Q000378:metabolism*; D020134:Catalytic Domain; D016825:Chlamydomonas reinhardtii / Q000201:enzymology*; D018360:Crystallography, X-Ray; D003545:Cysteine / Q000378:metabolism; D004220:Disulfides / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D013058:Mass Spectrometry; D009154:Mutation; D010084:Oxidation-Reduction / Q000187:drug effects; D010100:Oxygen / Q000378:metabolism; D045332:Photosystem II Protein Complex / Q000378:metabolism*; D017433:Protein Structure, Secondary
chemical_list: D002257:Carbonic Anhydrase Inhibitors; D004220:Disulfides; D045332:Photosystem II Protein Complex; D002256:Carbonic Anhydrases; D003545:Cysteine; D010100:Oxygen