The liver isoform of carnitine palmitoyltransferase 1 is not targeted to the endoplasmic reticulum
| Autor: | N. M. Broadway, E.David Saggerson, Nigel Turner, Richard J. Pease, Graeme M. Birdsey, Majid Shayeghi |
|---|---|
| Rok vydání: | 2003 |
| Předmět: |
Chloramphenicol O-Acetyltransferase
Male Transcription Genetic Mitochondrion Biology Endoplasmic Reticulum Biochemistry Rats Sprague-Dawley chemistry.chemical_compound Carnitine palmitoyltransferase 1 Animals Molecular Biology DNA Primers Microscopy Confocal Base Sequence Carnitine O-Palmitoyltransferase Endoplasmic reticulum food and beverages Cell Biology Peroxisome Molecular biology Fusion protein Rats Isoenzymes Malonyl-CoA chemistry Protein Biosynthesis COS Cells Porin Microsomes Liver Microsome Research Article |
| Zdroj: | Biochemical Journal. 370:223-231 |
| ISSN: | 1470-8728 0264-6021 |
| DOI: | 10.1042/bj20021269 |
| Popis: | Liver microsomal fractions contain a malonyl-CoA-inhibitable carnitine acyltransferase (CAT) activity. It has been proposed [Fraser, Corstorphine, Price and Zammit (1999) FEBS Lett. 446, 69—74] that this microsomal CAT activity is due to the liver form of carnitine palmitoyltransferase 1 (L-CPT1) being targeted to the endoplasmic reticulum (ER) membrane as well as to mitochondria, possibly by an N-terminal signal sequence [Cohen, Guillerault, Girard and Prip-Buus (2001) J. Biol. Chem. 276, 5403—5411]. COS-1 cells were transiently transfected to express a fusion protein in which enhanced green fluorescent protein was fused to the C-terminus of L-CPT1. Confocal microscopy showed that this fusion protein was localized to mitochondria, and possibly to peroxisomes, but not to the ER. cDNAs corresponding to truncated (amino acids 1—328) or full-length L-CPT1 were transcribed and translated in the presence of canine pancreatic microsomes. However, there was no evidence of authentic insertion of CPT1 into the ER membrane. Rat liver microsomal fractions purified by sucrose-density-gradient centrifugation contained an 88kDa protein (p88) which was recognized by an anti-L-CPT1 antibody and by 2,4-dinitrophenol-etomoxiryl-CoA, a covalent inhibitor of L-CPT1. Abundance of p88 and malonyl-CoA-inhibitable CAT activity were increased approx. 3-fold by starvation for 24h. Deoxycholate solubilized p88 and malonyl-CoA-inhibitable CAT activity from microsomes to approximately the same extent. The microsomal fraction contained porin, which, relative to total protein, was as abundant as in crude mitochondrial outer membranes fractions. It is concluded that L-CPT1 is not targeted to the ER membrane and that malonyl-CoA CAT in microsomal fractions is L-CPT1 that is derived from mitochondria, possibly from membrane contact sites. |
| Databáze: | OpenAIRE |
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