Plant Physiol.
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Plant Physiology 64:909-913 (1979)
© 1979 American Society of Plant Biologists

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Articles

Characterization of GDP-Fucose

Polysaccharide Fucosyl Transferase in Corn Roots (Zea mays L.) 1

Douglas W. James, Jr.2 and Russell L. Jones3

a Department of Botany, University of California, Berkeley, California 94720

The peripheral root cap cells of corn (cv. SX-17A) secrete a fucose-rich, high molecular weight, polysaccharide slime via the dictyosome pathway. To study the synthesis of this polysaccharide, a technique for isolating and assaying GDP-fucose:polysaccharide fucosyl transferase activity was developed. Corn roots were excised from germinated seeds, incubated 12 hours at 10 C in water, and ground in 100 millimolar Tris or Pipes buffer (pH 7.0) with or without 0.5 molar sucrose. The membrane-bound enzyme was solubilized by sonication in the presence of 2 molar urea and 1.5% (v/v) Triton X-100 and assayed by monitoring the incorporation of GDP-[14C]fucose into endogenous acceptors. Optimum enzyme activity is expressed at pH 7.0 and 30 C in the presence of 0.8% (v/v) Triton X-100. The enzyme does not require divalent cations for activation and is inhibited by concentrations of MnCl2 or MgCl2 greater than 1 millimolar. Corn root cap slime will serve as an exogenous acceptor for the enzyme if it is first hydrolyzed in 5 millimolar trifluoroacetic acid for 60 minutes at 18 pounds per square inch, 121 C. This procedure prepares the acceptor by removing terminal fucose residues from the slime molecule. Kinetics of fucose release during hydrolysis of native slime and in vitro synthesized product suggests that the two polymers possess similar linkages to fucose.

2 Present address: Department of Biochemistry, University of Texas Health Science Center, San Antonio, Texas 78284.

3 To whom reprint requests should be sent.

1 This research was supported by National Science Foundation Grant PCM 75-18870.






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ASPB Publications PLANT PHYSIOLOGY® THE PLANT CELL
Copyright © 1979 by the American Society of Plant Biologists