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Articles

Initial Organic Products of Assimilation of [¹³N]Ammonium and [¹³N]Nitrate by Tobacco Cells Cultured on Different Sources of Nitrogen ¹

W.-S. Chien

MSU-ERDA Plant Research Laboratory and Department of Physics, Michigan State University, East Lansing, Michigan 48824, Cyclotron Laboratory and Department of Physics, Michigan State University, East Lansing, Michigan 48824

Glutamine is the first major organic product of assimilation of ¹³NH₄⁺ by tobacco (Nicotiana tabacum L. cv. Xanthi) cells cultured on nitrate, urea, or ammonium succinate as the sole source of nitrogen, and of ¹³NO₃^– by tobacco cells cultured on nitrate. The percentage of organic ¹³N in glutamate, and subsequently, alanine, increases with increasing periods of assimilation. ¹³NO₃^–, used for the first time in a study of assimilation of nitrogen, was purified by new preparative techniques. During pulse-chase experiments, there is a decrease in the percentage of ¹³N in glutamine, and a concomitant increase in the percentage of ¹³N in glutamate and alanine. Methionine sulfoximine inhibits the incorporation of ¹³N from ¹³NH₄⁺ into glutamine more extensively than it inhibits the incorporation of ¹³N into glutamate, with cells grown on any of the three sources of nitrogen. Azaserine inhibits glutamate synthesis extensively when ¹³NH₄⁺ is fed to cells cultured on nitrate. These results indicate that the major route for assimilation of ¹³NH₄⁺ is the glutamine synthetase-glutamate synthase pathway, and that glutamate dehydrogenase also plays a role, but a minor one. Methionine sulfoximine inhibits the incorporation of ¹³N from ¹³NO₃^– into glutamate more strongly than it inhibits the incorporation of ¹³N into glutamine, suggesting that the assimilation of ¹³NH₄⁺ derived from ¹³NO₃^– may be mediated solely by the glutamine synthetase-glutamate synthase pathway.

³ Present address: Biology and Agriculture Division, Bhabha Atomic Research Centre, Trombay, Bombay 400 085, India.

⁴ Present address: Department of Bacteriology, University of California, Davis, California 95616.

¹ This work was supported by U.S. Energy Research and Development Administration Contract EY-76-C-02-1338 and by National Science Foundation Grants 74-01206 and 78-01684. This work is taken from a dissertation to be submitted to Michigan State University by T. A. S. in partial fulfillment of the requirements for the degree of Ph.D. in botany.