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Title: ALTERATION OF THE ADENINE NUCLEOTIDE RESPONSE AND INCREASED RUBISCO ACTIVATION ACTIVITY OF ARABIDOPSIS RUBISCO ACTIVASE BY SITE-DIRECTED MUTAGENESIS

Author
item KALLIS, RUSSELL - PLANT BIOLOGY UOFI URBANA
item EWY, ROBERT
item PORTIS JR, ARCHIE

Submitted to: Plant Physiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 4/4/2000
Publication Date: 7/1/2000
Citation: Kallis, R.P., Ewy, R.G., Portis Jr, A.R. 2000. Alteration of the adenine nucleotide response and increased rubisco activation activity of arabidopsis rubisco activase by side-directed mutagenesis. Plant Physiology. 123:1077-1086.

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. The activity of Rubisco is determined and hence regulated by another enzyme known as Rubisco activase. Rubisco activity might be increased to improve plant growth by altering its regulation. In this study, we found that mutations in a critical domain of the Rubisco activase protein increased its activity and altered its response to an important regulatory factor. 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: Arabidopsis ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) was activated in vitro at rates 2-3 fold greater by recombinant Arabidopsis 43-kD Rubisco activase with the amino acid replacements Q111E and Q111D in a phosphate binding loop, G-G-K-G-Q-G-K-S. However, these two mutants exhibited only slightly greater rates of ATP hydrolysis. Activities of Q11D mutant were much less sensitive and those of the Q111E mutant were somewhat less sensitive to inhibition by ADP. Both mutants exhibited higher Rubisco activation activities over they physiological range of ADP/ATP ratios. Mutants with non-polar, polar, and basic residues substituted at position Q111 exhibited rates of Rubisco activation less than the wildtype. Estimates of the relative affinity of the 43-kD wildtype and Q111D, E and S mutants for adenosine nucleotides by a variety of methods revealed that the affinities were the most diminished in the Q111D mutant, although the values obtained with each method varied considerably. The temperature stability of the Q111D and Q111E mutants did not differ markedly from the 43-kD recombinant wildtype, which is somewhat thermolabile. The Q111D and E mutants, expressed in planta, may provide a means to better define the role of ADP/ATP role in regulation of Rubisco activation and photosynthesis rate.