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Title: IDENTIFICATION OF CRITICAL ARGININE RESIDUES IN THE FUNCTIONING OF RUBISCO ACTIVASE

Author
item LI, CISHAN - UNIVERSITY OF ILLINIOS
item WANG, DAFU - UNIVERSITY OF ILLINOIS
item PORTIS JR, ARCHIE

Submitted to: Archives of Biochemistry and Biophysics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/30/2006
Publication Date: 5/5/2006
Citation: Li, C., Wang, D., Portis Jr, A.R. 2006. Identification of critical arginine residues in the functioning of Rubisco activase. Archives Of Biochemistry and Biophysics. 450:176-182.

Interpretive Summary: The activity of Rubisco, the enzyme that captures carbon dioxide, often limits photosynthesis, the process by which plants use light energy from the sun to make carbohydrates for growth from carbon dioxide and water. Rubisco activity is maintained by another chloroplast protein, Rubisco activase. Rubisco activity might be increased to improve plant growth by altering its interaction with Rubisco. The activase protein is a member of a large AAA+ protein family in which highly conserved arginine residues have been shown in a few family members to play critical roles in their functioning. In this work, we used site-directed mutagenesis of homologous arginine residues in activase to show that several of them are also critical in the functioning of activase. This information will benefit scientists attempting to modify the properties of Rubisco and the activase in ways beneficial for increased photosynthesis by crop plants.

Technical Abstract: Rubisco activase is a member of the AAA+ family in which arginines located in the Box VII and Sensor 2 domains are a recurrent feature and typically contribute to ATP-binding/hydrolysis or an inter-subunit interface. Replacement of R241 or R244 in Box VII or R294 or R296 in Sensor 2 with alanine in tobacco activase did not greatly alter the binding of ATP or ADP. However, ATP hydrolysis was minimal (R241A and R244A) or greatly diminished (R296A) and none of these mutants were able to activate Rubisco. R241, R244 and R296 were also required for nucleotide-dependent conformational changes detected by intrinsic fluorescence and limited proteolysis which may be associated with changes in oligomerization. However a nucleotide-dependent increase in the apparent molecular mass of a spinach R239A mutant occurred in the absence of both ATP hydrolysis activity and an intrinsic fluorescence increase which raises questions in interpreting previous correlations between these parameters.