THE CONVERSION OF ORNITHINE TO CITRULLINE BY RAT LIVER HOMOGENATES

The conversion of ornithine to citrulline as an obligatory step in the synthesis of urea in mammalian liver was first postulated by Krebs and Henseleit (1).This reaction was later investigated in detail by Gornall and Hunter (2) using rat liver slices.The use of liver slices for the study of this reaction is unsatisfactory in that citrulline, once formed from ornithine, is rapidly converted through arginine to urea.The use of homogenized liver preparations has eliminated this difficulty in part because of more adequate control of the conditions necessary for the various steps in the cycle.It was found early in this study that components and conditions necessary for the synthesis of citrulline from ornithine were such as to catalyze the disappearance of citrulline at an appreciable rate when magnesium ions were present, (3).The absence of magnesium ions, however, resulted in an accumulation of citrulline.This means of stopping the urea cycle at citrulline provided a convenient system for the study of the optimum conditions necessary for the conversion of ornithine to citrulline.Srb and Horowitz as a result of studies on Neurosporu mutants (4) have offered evidence for the existence of two distinct enzymatically catalyzed steps in the conversion of ornithine to citrulline, the first of these being the introduction of carbon dioxide, and the second, the introduction of ammonia.In the present work, this synthesis has been studied as a unit reaction.Experiments reported in this paper deal particularly with the formation of citrulline in the stepwise synthesis of urea from ornithine by rat liver homogenates.Procedures Preparation of Homogenates-A detailed description for the preparation of homogenates has been published previously (3).0.5 ml. of a 20 per cent rat liver homogenate prepared in isotonic KC1 was used in each incubation flask throughout the study, unless otherwise specified.This amount of homogenate contained between 2.3 and 2.8 mg. of tissue nitrogen as determined by micro-Kjeldahl analysis.