The isoenzyme pattern of chorismate mutase (EC 5.4.99.5) was examined by diethylaminoethyl-cellulose chromatography in a wide variety of plants. All plants contained a regulated form of chorismate mutase (CM-I), and most contained an additional, unregulated form (CM-2). The regulatory properties of CM-I differed significantly between plants. Antisera prepared against CM-I and CM-2 from Sorghum bicolor were used to test immunological cross reaction of chorismate mutases from other plants. There was a high degree of similarity between chorismate mutase isoenzymes from Sorghum bicolor and Zea mays and some with Hordeum vulgare, but all other species studied were antigenically distinct from sorghum. No homology between the structure of CM-I and CM-2 was detected within any species.The shikimate pathway provides the precursors for the biosynthesis of the aromatic amino acids phenylalanine, tyrosine, and tryptophan in plants as well as in microorganisms (8,12,19 (15).In spite of the importance of chorismate mutase in phenylalanine and tyrosine biosynthesis, the information available in the literature is limited to a very few species. In this report, we present the partial purification, the regulatory properties and antigenic similarity to the sorghum enzyme of chorismate mutases from several species, differing in morphology (monocots and dicots) and physiology (C3, C4, and CAM). MATERIALS AND METHODSPlant Material. The plants used for this study were selected on the basis of their morphological and physiological characteristics. Green seedlings of Avena sativa (C3, monocot), Brassica oleracea (C3, dicot), Hordeum vulgare (C3, monocot), Pennisetum typhoides (C4, monocot), and Zea mays (C4, monocot) were grown by planting seeds in wet vermiculite in a growth chamber at 28°C under 12 h fluorescent illumination. Shoots from 6-to 7-d-old seedlings were used for the extraction of enzyme. Young, green leaves of Amaranthus hypochondriacus (C4, dicot), Hoya carnosa (CAM, dicot), Medicago sativa (C3, dicot), and Xerosicyos danguyi (CAM, dicot) were obtained from adult plants grown in a greenhouse and exposed to normal spring and summer daylengths. Young, green leaves from wild adult plants of Eschscholtzia californica (C3, dicot) were used. Spinacia oleracea (C3, dicot) leaves were obtained from a local supermarket.Enzyme Extraction. For the extraction of chorismate mutase, 20 to 25 g of shoots or leaf tissues were powdered in liquid N2 and extracted at 0 to 4°C with one volume of buffer A (100 mM Tris-HCl, pH 8.0, containing 1 mM tryptophan and 0.1% 2-mercaptoethanol). The extract was filtered through two layers of cheesecloth (grade 60) and centrifuged at 60,000g for 90 min in a Beckman model L ultracentrifuge. All subsequent purification steps were carried out at 0 to 4°C. Desalting. The centrifuged supernatant was loaded at 7 ml/min onto a bed of Sephadex G-25 (2.5 x 29.0 717 www.plantphysiol.org on April 1, 2019 -Published by Downloaded from
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