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Activation of interspecies-hybrid Rubisco enzymes to assess different models for the Rubisco–Rubisco activase interaction

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Abstract

Ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) is prone to inactivation from non-productive binding of sugar-phosphates. Reactivation of Rubisco requires conformational remodeling by a specific chaperone, Rubisco activase. Rubisco activase from tobacco and other plants in the family Solanaceae is an inefficient activator of Rubisco from non-Solanaceae plants and from the green alga Chlamydomonas reinhardtii. To determine if the Rubisco small subunit plays a role in the interaction with Rubisco activase, a hybrid Rubisco (SSNT) composed of tobacco small subunits and Chlamydomonas large subunits was constructed. The SSNT hybrid, like other hybrid Rubiscos containing plant small subunits, supported photoautotrophic growth in Chlamydomonas, but growth in air was much slower than for cells containing wild-type Rubisco. The kinetic properties of the SSNT hybrid Rubisco were similar to the wild-type enzyme, indicating that the poor growth in air was probably caused by disruption of pyrenoid formation and the consequent impairment of the CO2concentrating mechanism. Recombinant Rubisco activase from Arabidopsis activated the SSNT hybrid Rubisco and hybrid Rubiscos containing spinach and Arabidopsis small subunits at rates similar to the rates with wild-type Rubisco. However, none of the hybrid Rubiscos was activated by tobacco Rubisco activase. That replacement of Chlamydomonas small subunits with plant small subunits does not affect the species-specific interaction between Rubisco and Rubisco activase suggests that the association is not dominated by the small subunits that surround the Rubisco central solvent channel. Therefore, the geometry of a side-on binding mode is more consistent with the data than a top-on or ring-stacking binding mode.

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Abbreviations

AAA+:

ATPase associated with diverse cellular activities

ER:

Rubisco in the uncarbamylated form and containing bound RuBP

ECM:

Rubisco carbamylated and containing bound Mg2+

K c :

K m for CO2

K o :

K m for O2

RuBP:

Ribulose 1,5-bisphosphate

Rubisco:

Ribulose-1,5-bisphosphate carboxylase/oxygenase

SSAT:

Hybrid Rubisco composed of Chlamydomonas large subunits and Arabidopsis small subunits

SSNT:

Hybrid Rubisco composed of Chlamydomonas large subunits and tobacco small subunits

SSSO:

Hybrid Rubisco composed of Chlamydomonas large subunits and spinach small subunits

V c :

V max of carboxylation

V o :

V max of oxygenation

Ω :

CO2/O2 specificity factor

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Acknowledgments

This study was funded by the Division of Chemical Sciences, Geosciences, and Biosciences, Office of Basic Energy Sciences, of the United States Department of Energy through Photosynthetic Systems Grants DE-FG02-00ER15044 to R.J.S., DE-FG02-09ER16123 to R.M.W. and DE-AI02-97ER20268 to M.E.S. The authors would like to thank Professor Govindjee for his many contributions to photosynthesis including encouraging publication of the first report of Rubisco activase in Photosynthesis Research 7: 193–201 (1985).

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Author notes

  1. A. Elizabete Carmo-Silva

    Present address: Plant Biology and Crop Science Department, Rothamsted Research, West Common, Harpenden, Herts, AL5 2JQ, UK

  2. Csengele Barta

    Present address: Department of Biology, Missouri Western State University, 4525 Downs Drive, Agenstein Hall 237G, Saint Joseph, MO, 64507, USA

  3. Todor Genkov

    Present address: Center for Biocatalysis and Bioprocessing, University of Iowa, Coralville, IA, 52241, USA

Authors and Affiliations

  1. Department of Chemistry and Biochemistry, Arizona State University, Tempe, AZ, 85287, USA

    Rebekka M. Wachter

  2. US Arid-Land Agricultural Research Center, Agricultural Research Service, U.S. Department of Agriculture (USDA-ARS), 21881 N. Cardon Lane, Maricopa, AZ, 85138, USA

    Michael E. Salvucci, A. Elizabete Carmo-Silva & Csengele Barta

  3. Department of Biochemistry, University of Nebraska, Lincoln, NE, 68588, USA

    Todor Genkov & Robert J. Spreitzer

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  1. Rebekka M. Wachter
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Correspondence to Michael E. Salvucci.

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Wachter, R.M., Salvucci, M.E., Carmo-Silva, A.E. et al. Activation of interspecies-hybrid Rubisco enzymes to assess different models for the Rubisco–Rubisco activase interaction. Photosynth Res 117, 557–566 (2013). https://doi.org/10.1007/s11120-013-9827-0

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