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Title: SPECIFICITY OF ACTIVASE IS CHANGED BY A PRO-89 TO ARG SUBSTITUTION IN THE LARGE SUBUNIT OF RIBULOSE-1,5-BISPHOSPHATE CARBOXYLASE/OXYGENASE

Author
item LARSON, ERIC - CROP SCIENCES UI URB IL
item O'BRIEN, CAROLYN - BIOCHEMISTRY UNIV NEB
item ZHU, GENHAI - BIOCHEMISTRY UNIV NEB
item SPREITZER, ROBERT - BIOCHEMISTRY UNIV NEB
item PORTIS JR, ARCHIE

Submitted to: Journal of Biological Chemistry
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/9/1997
Publication Date: N/A
Citation: N/A

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. In order to have activity, Rubisco requires modification by a process called activation which is controlled by another protein known as Rubisco activase. Rubisco activity might be increased to improve plant growth by altering the regulation of activation. Activation by Rubisco activase is believed to involve binding of the proteins to each other because the proteins from one species of plants, the Solanaceae, do not work in combination with either protein from other species. The Rubisco protein in these plants has several unique amino acid differences located on the protein surface that may determine the nature of the interaction with Rubisco activase. In this study we found that changing only one of these amino acids was sufficient to allow Rubisco from a non-Solanaceous species to be activated by a Solanaceous Rubisco activase, but not by the non-Solanaceous protein that worked before this change. This information will benefit scientists attempting to modify the properties and regulation of Rubisco in ways beneficial for increased photosynthesis by crop plants.

Technical Abstract: Tobacco activase does not markedly facilitate the activation of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco, EC 4.1.1.39) from non-Solanaceae species, including the green alga Chlamydomonas reinhardtii. To examine the basis of this specificity, we focused on two exposed residues in the large subunit of Rubisco that are unique to the Solananceae proteins. By employing in vitro mutagenesis and chloroplast transformation, P89R and K356Q substitutions were separately made in the Chlamydomonas enzyme to change these residues to those present in tobacco. Both mutants were indistinguishable from the wild-type when grown with minimal medium in the light and contained wild-type levels of holoenzyme. Purified Rubisco was assessed for facilitated activation by spinach and tobacco activase. Both wild-type and K356Q Rubisco were similar in that spinach activase was much more effective than tobacco activase. In contrast, P89R Rubisco was not activated by spinach activase, but was activated well by tobacco activase. Thus, the relative specificities of the spinach and tobacco activases for Chlamydomonas Rubisco were switched by changing a single residue at position 89. This result provides evidence for a site on the Rubisco holoenzyme that interacts directly with Rubisco activase.