Journal Article

A Dithiothreitol-Sensitive Tetrameric Protease from Spinach Thylakoids Has Polyphenol Oxidase Activity

Tomohiko Kuwabara, Tetsuya Masuda and Sachiko Aizawa

in Plant and Cell Physiology

Published on behalf of Japanese Society of Plant Physiologists

Volume 38, issue 2, pages 179-187
Published in print January 1997 | ISSN: 0032-0781
Published online January 1997 | e-ISSN: 1471-9053 | DOI: http://dx.doi.org/10.1093/oxfordjournals.pcp.a029150
A Dithiothreitol-Sensitive Tetrameric Protease from Spinach Thylakoids Has Polyphenol Oxidase Activity

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Polyclonal antibody raised against a dithiothreitol-sen-sitive tetrameric protease (DSTP) from PSII membranes specifically inhibited the polyphenol oxidase (PPO) activity of spinach thylakoids. DSTP was copurified with PPO activity on an affinity column prepared with antibody against DSTP. These results suggest that DSTP and PPO are the same protein. During purification of DSTP, Tween 20 was essential for stabilization of the protein, which was degraded in the absence of the detergent. Gel-filtration chromatog-raphy of the purified DSTP revealed the presence of 230-kDa (tetramer) and 60-kDa (monomer) species. The copper content of monomer species was determined to be 0.4 Cu atom per protein molecule, when the molecular weight of the protein was calculated to be 62,243, which is the value reported for spinach PPO [Hind et al. (1995) Biochemistry 34: 8157]. Purified DSTP caused the degradation as well as the dimerization of the extrinsic 23-kDa protein of PSII. The degradation of the protein was suppressed under anaerobic conditions induced by the presence of glucose oxidase and glucose together. This fact suggests that oxygen molecules are involved in the proteolytic reaction and that the proteolytic activity and PPO activity may be correlated with each other.

Keywords: Active oxygen; Copper; Dithiothreitol-sen-sitive tetrameric protease; Photosystem II; Polyphenol oxidase (EC 1.10.3.1); Proteolysis

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Subjects: Biochemistry ; Molecular and Cell Biology ; Plant Sciences and Forestry

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