Book chapter
[16] Actin amino-terminal acetylation and processing in a rabbit reticulocyte lysate
Methods in Enzymology, pp.179-192
Elsevier Science & Technology
1984
DOI: 10.1016/0076-6879(84)06018-3
PMID: 6493055
Abstract
This chapter describes method for inhibiting the acetylation of proteins in the reticulocyte lysate. It also illustrates procedure for assaying the acetylation of the actin NH2-terminal methionine residue and the subsequent removal of acetylmethionine from the completed actin polypeptide chain. A nonreactive analog of acetyl-CoA, S-acetonyl-CoA, is synthesized to serve as a potential competitive inhibitor of the protein acetyltransferase. If the reticulocyte lysate is first treated according to Palmiter and then S-acetonyl-CoA is added, then 85-95% inhibition of acetylation could be achieved. It has been demonstrated that NH2-terminal acetylation can occur as a cotranslational process after the first 30-40 amino acids of the nascent polypeptide chain have been polymerized. The chapter considers using acetylation inhibition system to make actin as a completed 43,000-dalton polypeptide with a free NH2- terminal methionine. Following this, the acetylation of the NH2-terminal methionine can occur in a fully posttranslational acetyl- CoA-dependent reaction in the reticulocyte lysate.
Details
- Title: Subtitle
- [16] Actin amino-terminal acetylation and processing in a rabbit reticulocyte lysate
- Creators
- Kent L RedmanPeter A Rubenstein
- Resource Type
- Book chapter
- Publication Details
- Methods in Enzymology, pp.179-192
- Publisher
- Elsevier Science & Technology
- DOI
- 10.1016/0076-6879(84)06018-3
- PMID
- 6493055
- eISSN
- 1557-7988
- ISSN
- 0076-6879
- Language
- English
- Date published
- 1984
- Academic Unit
- Stead Family Department of Pediatrics; Biochemistry and Molecular Biology; Internal Medicine
- Record Identifier
- 9984024563402771
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