Plant Physiol. Journal of Pharmacology and Experimental Therapeutics
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Plant Physiology 78:221-227 (1985)
© 1985 American Society of Plant Biologists

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Articles

L-Aspartate Transport into Pea Chloroplasts 1

Kinetic and Inhibitor Evidence for Multiple Transport Systems

Margaret Werner-Washburne and Kenneth Keegstra

Department of Botany, University of Wisconsin, Madison, Wisconsin 53706

The kinetics of L-aspartate transport into pea chloroplasts was studied in the presence and absence of transport inhibitors to determine whether multiple aspartate carriers exist. Transport was measured by the silicone oil centrifugation technique. Reciprocal plots of concentration-dependent transport rates were biphasic, indicating the presence of two transport components, distinguishable on the basis of their affinity for aspartate. These transport components, called high affinity and low affinity transport could also be distinguished on the basis of their apparent substrate saturability and their sensitivity to media pH. The apparent Km for high affinity transport was 30 micromolar. The Km for low affinity transport was not determined. To test whether these transport components could also be distinguished on the basis of inhibitor sensitivity and to assess the value of inhibitors for distinguishing multiple aspartate translocators, a survey of several classes of potential inhibitors was conducted. High affinity aspartate transport was inhibited by p-chloromercuribenzenesulfonate and mersalyl, both sulfhydryl-reactive reagents; diethyl pyrocarbonate, a histidine-reactive reagent; and nigericin and carbonyl cyanide m-chlorophenylhydrazone, both ionophores. Low affinity aspartate transport was not inhibited by p-chloromercuribenzenesulfonate or nigericin, but preliminary results suggest it was sensitive to diethyl pyrocarbonate. Because the high and low affinity transport components could be distinguished not only by their sensitivity to media pH and substrate saturability, but also by their sensitivity to various inhibitors, we concluded that they may represent different transport systems or carriers.

1 Supported in part by a grant from the United States Department of Agriculture Competitive Research Office. M. W.-W. was supported by National Institutes of Health Grant 5T32 GM 07215-07.






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Copyright © 1985 by the American Society of Plant Biologists