Abstract

Rubisco (ribulose-1,5-bisphosphate carboxylase/oxygenase) catalyses CO2 assimilation in biology. A prerequisite for catalysis is an activation pro- cess, whereby an active site lysine is selectively carbamylated. The carba- myl group is then stablised by a metal ion, which in vivo is Mg2+. Other divalent metal ions can replace Mg2+ as activators in vitro, but the nature of the metal ion strongly influences the catalytic activity of the enzyme and has a differential effect on the ratio of the carboxylation reaction and the competing oxygenation reaction. Biochemical studies show that calcium promotes carbamylation but not catalysis. To investigate the role of the metal in catalysis, we have determined two structures of the enzyme complexed with Ca2+ and the transition state analogue 2-carboxy-D-arbinitol-1,5-bisphosphate (2CABP). One of the complexes was prepared by soaking 2CABP into crystals of the enzyme-Ca2+-product complex, while the other was obtained by cocrystallising the enzyme with calcium and 2CABP under activating conditions. The two crystals belong to different space groups, and one was merohedrally twinned. Both complexes show very similar three-dimensional features. The enzyme is carbamylated at Lys201, and requisite loops close over the bound ligands in the active site, shielding them from the solvent in a manner similar to the corresponding complex with Mg2+. However, there are subtle differences that could explain the particular role of Ca2+ in these processes. The larger radius of the calcium ion and its reduced Lewis-acid character causes a significant increase in the required proton hop distance between the C3 proton and the carbarnate on Lys201 in the calcium complex. This alone could explain the inability of calcium to sustain catalysis in Rubisco. Similar effects are also expected on subsequent proton transfer steps in the catalytic cycle. Here we also discuss the effect of metal substitution on the dynamics of the ligands around the metal ion. (C) 2003 Elsevier Ltd. All rights reserved

Keywords

Rubisco; Calcium activation; Loop movements; Ligand complexes; merohedral twinning

Published in

Journal of Molecular Biology
2003, Volume: 334, number: 1, pages: 65-73
Publisher: ACADEMIC PRESS LTD ELSEVIER SCIENCE LTD

SLU Authors

• Karkehabadi, Saeid

  • Department of Molecular Biology, Swedish University of Agricultural Sciences
    

• Andersson, Inger

  • Department of Molecular Biology, Swedish University of Agricultural Sciences
    

• Taylor, Tom

  • Department of Molecular Biology, Swedish University of Agricultural Sciences
    

UKÄ Subject classification

Forest Science
Food Science
Environmental Sciences related to Agriculture and Land-use


Publication identifier

DOI: https://doi.org/10.1016/j.jmb.2003.09.025

Permanent link to this page (URI)

https://res.slu.se/id/publ/2461