HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Sharma, C. P. Articles by Gehring, H. Search for Related Content PubMed PubMed Citation Articles by Sharma, C. P. Articles by Gehring, H. Social Bookmarking                      What's this?

J. Biol. Chem., Vol. 261, Issue 24, 11146-11149, Aug, 1986

The precursor of mitochondrial aspartate aminotransferase is translocated into mitochondria as apoprotein

CP Sharma and H Gehring

Mitochondrial aspartate aminotransferase is synthesized on free polysomes as a higher molecular weight precursor (Sonderegger, P., Jaussi, R., Christen, P., and Gehring, H. (1982) J. Biol. Chem. 257, 3339-3345). The present study examines whether the coenzyme pyridoxal phosphate or pyridoxamine phosphate is required for the uptake of the precursor into mitochondria. Chicken embryo fibroblasts were cultured in medium prepared with and without pyridoxal. In cells grown in the presence of pyridoxal only holoform of aspartate aminotransferase and no apoenzyme was detected. Cells cultured under pyridoxal deficiency contained about 30% of apoenzyme in secondary cultures. All of this apoform was identified as mitochondrial isoenzyme. In order to differentiate whether this apoenzyme corresponded to newly synthesized protein or originated from pre-existing holoenzyme, double isotope- labeling experiments were performed. Secondary cultures of chicken embryo fibroblasts grown under pyridoxal depletion were labeled with [3H]methionine, and then pulsed with [35S]methionine. In another series of experiments, the 3H-labeled cells were pulsed with [35S]methionine in the presence of the protonophore carbonyl cyanide m- chlorophenylhydrazone in order to accumulate the precursor. Subsequently, the accumulated precursor was chased into the mitochondria by addition of the carbonyl cyanide m- chlorophenylhydrazone antagonist cysteamine. The holo- and apoenzyme from the ultrasonic extract of the double-labeled cells were separated by affinity chromatography on a phosphopyridoxyl-AH-Sepharose column, immunoprecipitated, and analyzed by sodium dodecyl sulfate- polyacrylamide gel electrophoresis and fluorography. Under both experimental conditions, the 3H/35S ratio of the apoenzyme was less than half of that of the holoenzyme. Therefore, the apoenzyme and not the holoenzyme is the first product of the precursor in the mitochondria. Apparently, the precursor of mitochondrial aspartate aminotransferase is transported into mitochondria as apoprotein and is processed there independently of the coenzyme.
CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				K. M. Robinson and B. D. Lemire Covalent Attachment of FAD to the Yeast Succinate Dehydrogenase Flavoprotein Requires Import into Mitochondria, Presequence Removal, and Folding J. Biol. Chem., February 23, 1996; 271(8): 4055 - 4060. [Abstract] [Full Text] [PDF]