Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: May 15, 2008
Publication Date: June 27, 2008
Citation: Hatfield, R.D., Sullivan, M.L., Schmitz, G. 2008. Characteristics of polyphenol oxidases [abstract]. The 2008 Joint Meeting of the American Society of Plant Biologists and the Sociedad Mexicana de Bioquimica Rama: Bioquimica y Biologia Molecular de Plantas, June 26-July 1, 2008, Merida, Mexico. Abstract P04010. p. 103. Technical Abstract: Polyphenol oxidase (PPO, EC 18.104.22.168 or EC 22.214.171.124) catalyzes the oxidation of o-diphenols to o-quinones. Highly reactive o-quinones couple with phenolics and specific amino acids on proteins to form the characteristic browning products in many wounded fruits, vegetables, and leaf tissues of plants like red clover and tomato. Production of o-quinones in red clover inhibits post-harvest proteolysis during the ensiling process. We established a model system to study PPO characteristics by expressing red clover PPO cDNA in alfalfa, which normally lacks foliar PPO activity and o-diphenols. Activation of PPO occurs upon incubation at room temperature for several days (4-10 days), resulting in a 40-fold increase in activity. Proteolytic activity against PPO appears to aid in its transition to a more active form, but is not solely dependent upon it. RuBisCo subunits were isolated from alfalfa and used as a model protein to study the interaction of PPO-generated o-quinones with proteins. Treatment of RuBisCo with PPO and caffeic or chlorogenic acid as o-diphenol substrates resulted in decreased migration of protein bands on SDS page gels. This would implicate PPO-generated o-quinones as possible cross-linking agents of proteins. However, digestion of PPO/caffeic or chlorogenic-treated protein with trypsin did not inhibit RuBisCo's degradation, based on band migration on SDS-PAGE gels. Closer examination by peptide analysis may reveal minor shifts in the protein degradation patterns and resulting small peptides. Knowledge of red clover PPOs may improve our understanding of their in vivo function(s), and how their roles in post-harvest physiology may be exploited.