Coenzyme site-directed mutants of photosynthetic A4-GAPDH show selectively reduced NADPH-dependent catalysis, similar to regulatory AB-GAPDH inhibited by oxidized thioredoxin

Journal of Molecular Biology
Francesca SparlaPaolo Trost

Abstract

Chloroplast glyceraldehyde-3-phosphate dehydrogenase (GAPDH) of higher plants uses both NADP(H) and NAD(H) as coenzyme and consists of one (GapA) or two types of subunits (GapA, GapB). AB-GAPDH is regulated in vivo through the action of thioredoxin and metabolites, showing higher kinetic preference for NADPH in the light than in darkness due to a specific effect on kcat(NADPH). Previous crystallographic studies on spinach chloroplast A4-GAPDH complexed with NADP or NAD showed that residues Thr33 and Ser188 are involved in NADP over NAD selectivity by interacting with the 2'-phosphate group of NADP. This suggested a possible involvement of these residues in the regulatory mechanism. Mutants of recombinant spinach GapA (A4-GAPDH) with Thr33 or Ser188 replaced by Ala (T33A, S188A and double mutant T33A/S188A) were produced, expressed in Escherichia coli, and compared to wild-type recombinant A4-GAPDH, in terms of crystal structures and kinetic properties. Affinity for NADPH was decreased significantly in all mutants, and kcat(NADPH) was lowered in mutants carrying the substitution of Ser188. NADH-dependent activity was unaffected. The decrease of kcat/Km of the NADPH-dependent reaction in Ser188 mutants resembles the behaviour of ...Continue Reading

References

Mar 1, 1991·Acta Crystallographica. Section A, Foundations of Crystallography·T A JonesM Kjeldgaard
Oct 3, 1998·Acta Crystallographica. Section D, Biological Crystallography·A T BrüngerG L Warren
Oct 8, 1998·Acta Crystallographica. Section D, Biological Crystallography·S SongZ Lin
Mar 25, 1999·Acta Crystallographica. Section D, Biological Crystallography·P SabatinoP Trost
Dec 14, 2001·The Journal of Biological Chemistry·Fabrice MoucheNicolas Boisset
Nov 9, 2002·European Journal of Biochemistry·Renate ScheibeSonja-Manuela Sauermann
Jul 9, 2003·Biochemistry·Emmanuelle GracietBrigitte Gontero
Sep 1, 1994·Acta Crystallographica. Section D, Biological Crystallography·UNKNOWN Collaborative Computational Project, Number 4
Jan 1, 1997·Methods in Enzymology·E A Merritt, D J Bacon
Jan 1, 1997·Methods in Enzymology·Zbyszek Otwinowski, Wladek Minor

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Citations

Apr 2, 2008·Antioxidants & Redox Signaling·Peter Schürmann, Bob B Buchanan
Aug 2, 2005·Plant Physiology·Francesca SparlaPaolo Trost
Apr 4, 2006·Acta Crystallographica. Section F, Structural Biology and Crystallization Communications·Tomoya KitataniToshiji Tada
Aug 2, 2006·Acta Crystallographica. Section F, Structural Biology and Crystallization Communications·Tomoya KitataniToshiji Tada
Nov 2, 2006·Acta Crystallographica. Section F, Structural Biology and Crystallization Communications·Ana Cámara-ArtigasJames P Allen
Jun 3, 2010·Acta Crystallographica. Section F, Structural Biology and Crystallization Communications·Simona FermaniPaolo Trost
Jun 19, 2007·Proceedings of the National Academy of Sciences of the United States of America·S FermaniP Trost
Feb 14, 2012·Plant Science : an International Journal of Experimental Plant Biology·Mirko ZaffagniniPaolo Trost
Jun 5, 2015·Frontiers in Molecular Biosciences·Gabriel Thieulin-PardoBrigitte Gontero
Sep 10, 2014·The Journal of Biological Chemistry·Samuel MorisseStéphane D Lemaire
Jul 8, 2020·International Journal of Molecular Sciences·Igor PokotyloEric Ruelland
Jan 27, 2021·Antioxidants·Anna DreyerKarl-Josef Dietz
Apr 25, 2021·Trends in Plant Science·Libero GurrieriPaolo Trost

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