Uptake and Processing of Serine:Pyruvate Aminotransferase Precursor by Rat Liver Mitochondria In Vitro and In Vivo1

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Abstract
Processing and uptake of the precursor of serine: pyruvate aminotransferase [EC 2.6.1.51] by mitochondria were studied in vitro and in vivo . Serine: pyruvate aminotransferase was synthesized mainly on free ribosomes as judged by immunoprecipitation of puromycin-labeled nascent peptides prepared from free and bound ribosomes. The precursor of rat liver serine: pyruvate aminotransferase (pSPT) synthesized in vitro was post-translationally processed to an apparently mature form by isolated rat liver mitochondria. Available evidence indicated that the processed product was localized in the matrix of mitochondria. Mature serine: pyruvate aminotransferase did not inhibit the in vitro processing, suggesting that the extra peptide was necessary for the mitochondrial uptake of the precursor. In the livers of rats fed a vitamin B 6 -deficient high-protein diet, the induction by glucagon of serine: pyruvate aminotransferase occurred and most of the induced enzyme existed in mitochondria as the apo-form, suggesting that pSPT was taken up by mitochondria and processed in the apo-form under the conditions employed. In the in vitro system, on the other hand, the processing of pSPT proceeded both in the absence and presence of pyridoxal 5'-phosphate. Should the precursor also bind the prosthetic molecule, therefore, it would be transported into mitochondria in both the apo- and holo-forms. When isolated rat hepatocytes were labeled with [ 35 S]methionine, labeled pSPT appeared in the cytosolic fraction and was transported rapidly into mitochondria in association with the processing. This uptake and processing were inhibited by a fluorescent laser dye, rhodamine 123, and the precursor accumulated in the cytosol in the presence of the dye.

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