Abstract
The cationic isozyme of peroxidase isolated from suspension cultures of peanut cells is a heme-containing and calcium-dependent glycoprotein having four covalently attached oligosaccharide chains. Attempts were made to crystallize the glycoprotein for X-ray diffraction analysis, and these have met with some success. Crystals have now been grown that are suitable for a full three-dimensional structural analysis. The crystals are thin plates and we have shown them to be of the orthorhombic space group P2(1)2(1)2(1) with a = 48.1, b = 97.2, c = 146.2 A. The crystals diffract to beyond 2.8 A resolution, appear to be stable to lengthy X-ray exposure, and contain two molecules of 40,000 daltons each in the asymmetric unit.
Highlights
Most higher plants produce a variety of isozymic forms of the enzyme peroxidase (E.C. 1.11.1.7) which has been used as a convenient marker in genetic, physiological and pathological studies (Greppin, Penel & Gaspar, 1986; van Huystee, 1987)
The major cationic isozyme of cultured peanut cells was isolated and purified to homogeneity from the culture medium according to the procedures of Hu, Krol & van Huystee (1990)
Rather few glycoproteins having a percentage of carbohydrate as high as 20% have been crystallized, and those that have, in general, produced diffraction patterns of quite limited extent
Summary
Most higher plants produce a variety of isozymic forms of the enzyme peroxidase (E.C. 1.11.1.7) which has been used as a convenient marker in genetic, physiological and pathological studies (Greppin, Penel & Gaspar, 1986; van Huystee, 1987). Peroxidases are synthesized by cultured plant cells which secrete the cationic isozyme into the medium. It provides, a straightforward means for its purification (van Huystee, 1987; Stephan & van Huystee, 1981). The major cationic isozyme of peanut cell peroxidase has a total molecular weight of 40 000 daltons It consists of a single polypeptide chain 307 residues in length of 32 954 daltons molecular weight. This protein component is covalently attached to four polysaccharide chains of total weight 8500 daltons comprising 21% of the total glycoprotein molecular weight. We report below our first steps in this direction which we hope may provide a sound structural basis for the interpretation and correlation of its many biochemical features
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