|Stipanovic, Robert - Bob|
Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: 7/9/2006
Publication Date: 9/7/2006
Citation: Veshkurova, O., Arzanova, I.A., Ibragimov, F.A., Golubenko, Z., Akhunov, A.A., Salikhov, S.I., Stipanovic, R.D. 2006. Identification of IAA-oxidase in peroxidase isozymes from cotton plant leaves [abstract]. In: Proceedings of Peroxidase 2006, July 7-9, 2006, Aveiro, Portugal. p. 44.
Technical Abstract: One of the functions of plant peroxidase is to regulate the indole acetic acid (IAA) hormonal level by oxidizing it to inactive 3-methyleneoxyindole. IAA-binding proteins and plant peroxidase revealed five structurally similar fragments. We have isolated peroxidase isozymes with IAA-oxidase activity using affinity chromatography. Thus, a biospecific absorbent was synthesized for IAA-oxidase by conjugating IAA to AH-agarose. A protein fraction that did not bind to the affinity column had a specific peroxidase activity, 0.01 ncat/mg). The binding proteins were eluted by NaCl (1 M). The resulting eluate was desalted by dialysis and lyophilized (specific peroxidase activity, 0.091 ncat/mg). Next, binding constants of 3**H-IAA to peroxidase and the cotton plant auxin receptor that was isolated by our previous work were determined and compared. The binding was performed in the presence of increasing concentrations of unlabeled IAA (from 10**-9 to 10**-3 M). We found that 3**H-IAA is bound specifically with saturation to peroxidase from cotton leaves. However, the binding strength is three orders of magnitude less than to the auxin receptor (Kd 2x10**-6 and 6x19**-9 M, respectively). Adding a 1000-fold excess of the specific substrates benzidine or synthetic auxin alpha-naphthylacetic acid (NAA) to the analyzed sample inhibits the enzyme and partially inhibits 3**H-IAA binding. The results indicate that peroxidase is not a true auxin receptor. Auxin binding by the recognition segment of the enzyme may be required for the correct orientation of themolecule during its oxidation at the catalytic center.