Kim, Hyun Soo and Hoja, Ursula and Stolz, Juergen and Sauer, Guido and Schweizer, Eckhart (2004) Identification of the tRNA-binding protein Arc1p as a novel target of in vivo biotinylation in Saccharomyces cerevisiae. JOURNAL OF BIOLOGICAL CHEMISTRY, 279 (41). pp. 42445-42452. ISSN 0021-9258, 1083-351X
Full text not available from this repository. (Request a copy)Abstract
Biotin is an essential cofactor of cell metabolism serving as a protein-bound coenzyme in ATP-dependent carboxylation, in transcarboxylation, and certain decarboxylation reactions. The involvement of biotinylated proteins in other cellular functions has been suggested occasionally, but available data on this are limited. In the present study, a Saccharomyces cerevisiae protein was identified that reacts with streptavidin on Western blots and is not identical to one of the known biotinylated yeast proteins. After affinity purification on monomeric avidin, the biotinylated protein was identified as Arc1p. Using C-14-labeled biotin, the cofactor was shown to be incorporated into Arc1p by covalent and alkali-stable linkage. Similar to the known carboxylases, Arc1p biotinylation is mediated by the yeast biotin: protein ligase, Bpl1p. Mutational studies revealed that biotinylation occurs at lysine 86 within the N-terminal domain of Arc1p. In contrast to the known carboxylases, however, in vitro biotinylation of Arc1p is incomplete and increases with BPL1 overexpression. In accordance to this fact, Arc1p lacks the canonical consensus sequence of known biotin binding domains, and the bacterial biotin: protein ligase, BirA, is unable to use Arc1p as a substrate. Arc1p was shown previously to organize the association of MetRS and GluRS tRNA synthetases with their cognate tRNAs thereby increasing the substrate affinity and catalytic efficiency of these enzymes. Remarkably, not only biotinylated but also the biotin-free Arc1p obtained by replacement of lysine 86 with arginine were capable of restoring Arc1p function in both arc1Delta and arc1Deltalos1Delta mutants, indicating that biotinylation of Arc1p is not essential for activity.
| Item Type: | Article |
|---|---|
| Uncontrolled Keywords: | ACETYL-COA-CARBOXYLASE; FATTY-ACID SYNTHESIS; BIOTIN HOLOENZYME SYNTHETASE; ESCHERICHIA-COLI; RECOMBINANT PROTEINS; POSTTRANSLATIONAL MODIFICATION; ENZYMATIC BIOTINYLATION; YEAST; GENE; COENZYME; |
| Subjects: | 500 Science > 580 Botanical sciences |
| Divisions: | Biology, Preclinical Medicine > Institut für Pflanzenwissenschaften > Lehrstuhl für Zellbiologie und Pflanzenphysiologie (Prof. Dr. Klaus Grasser) |
| Depositing User: | Dr. Gernot Deinzer |
| Date Deposited: | 28 Jun 2021 05:12 |
| Last Modified: | 28 Jun 2021 05:12 |
| URI: | https://pred.uni-regensburg.de/id/eprint/37086 |
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