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Journal of Bacteriology, June 2005, p. 4222-4228, Vol. 187, No. 12
0021-9193/05/$08.00+0     doi:10.1128/JB.187.12.4222-4228.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Mutagenesis at Two Distinct Phosphate-Binding Sites Unravels Their Differential Roles in Regulation of Rubisco Activation and Catalysis

Yehouda Marcus,* Hagit Altman-Gueta, Aliza Finkler, and Michael Gurevitz*

Department of Plant Sciences, George S. Wise Faculty of Life Sciences, Tel-Aviv University, Ramat-Aviv 69978, Tel-Aviv, Israel

Received 16 November 2004/ Accepted 4 March 2005

Orthophosphate (Pi) has two antagonistic effects on ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco), stimulation of activation and inhibition of catalysis by competition with the substrate RuBP. The enzyme binds Pi at three distinct sites, two within the catalytic site (where 1P and 5P of ribulose 1,5-bisphosphate [RuBP] bind), and the third at the latch site (a positively charged pocket involved in active-site closure during catalysis). We examined the role of the latch and 5P sites in regulation of Rubisco activation and catalysis by introducing specific mutations in the enzyme of the cyanobacterium Synechocystis sp. strain PCC 6803. Whereas mutations at both sites abolished the Pi-stimulated Rubisco activation, substitution of residues at the 5P site, but not at the latch site, affected the Pi inhibition of Rubisco catalysis. Although some of these mutations substantially reduced the catalytic turnover of Rubisco and increased the Km(RuBP), they had little to moderate effect on the rate of photosynthesis and no effect on photoautotrophic growth. These findings suggest that in cyanobacteria, Rubisco does not limit photosynthesis to the extent previously estimated. These results indicate that both the latch and 5P sites participate in regulation of Rubisco activation, whereas Pi binding only at the 5P site inhibits catalysis in a competitive manner.


* Corresponding author. Mailing address: Department of Plant Sciences, George S. Wise Faculty of Life Sciences, Tel-Aviv University, Ramat-Aviv 69978, Tel-Aviv, Israel. Phone: 972-3-6406101 (Y. Marcus); 972-3-6409844 (M. Gurevitz). Fax: 972-3-6406100. E-mail: yehoudam{at}post.tau.ac.il; mamgur{at}post.tau.ac.il.


Journal of Bacteriology, June 2005, p. 4222-4228, Vol. 187, No. 12
0021-9193/05/$08.00+0     doi:10.1128/JB.187.12.4222-4228.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.




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