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United States Department of Agriculture

Agricultural Research Service

Research Project: IDENTIFYING AND MANIPULATING DETERMINANTS OF PHOTOSYNTHATE PRODUCTION AND PARTITIONING

Location: Global Change and Photosynthesis Research Unit

Title: A Spectrophotometric Assay to Measure Rubisco Activase Activation Activity under Varying Atp:adp Ratios

Authors
item Schrader, Stephen
item Salvucci, Michael
item Portis Jr, Archie

Submitted to: American Society of Plant Biologists
Publication Type: Abstract Only
Publication Acceptance Date: June 1, 2006
Publication Date: August 5, 2006
Citation: Schrader, S.M., Salvucci, M.E., Portis Jr, A.R. 2006. A spectrophotometric assay to measure rubisco activase activation activity under varying ATP:ADP ratios [abstract]. American Society of Plant Biologists. Paper No. P14008.

Technical Abstract: The ratio of ATP to ADP in the stroma is an important regulatory mechanism for controlling the activation state of Rubisco via Rubisco activase (activase). Understanding the response of activase to a varying ATP:ADP ratio should reveal insights into the regulation of photosynthesis. However, the current spectrophotometric method for measuring Rubsico activity requires saturating ATP for the coupling enzymes to function properly. This limits the measurement of activase activation activity to radiometric methods, which can be costly and less safe than spectrophotometric methods. We report here on a spectrophotometric assay that measures Rubisco activation by activase using phosphoglycerate mutase, enolase, pyruvate kinase, and lactic dehydrogenase as coupling enzymes. This new assay has equivalent results to the traditional Rubisco spectrophotometric assay at 25C without the need for saturating ATP. The assay is also stable at 45C with 10 'g mL-1 of Rubisco. Higher Rubisco concentrations had lower activities at 45C when using 10 U mL-1 of coupling enzymes. Rubisco activation results by activase will be presented.

Last Modified: 8/27/2014