In Vitro Synthesis of pp60v-src: Myristylation in a Cell-Free System

• DOI: 10.1128/mcb.8.10.4295-4301.1988
• Publication Date: 2021-05-28T14:33:20Z
• Authors: Ida Deichaite; Lawrence P. Casson; Huai-Ping Ling; Marilyn D. Resh
• Keywords: Rous sarcoma virus; Reticulocyte; Cell-free system

Covalent attachment of myristic acid to pp60v-src, the transforming protein Rous sarcoma virus, was studied in a cell-free system. Using synthetic peptide containing first 11 amino acids mature pp60v-src polypeptide sequence as substrate, we probed lysates from variety cells and tissues for N-myristyl transferase (NMT) activity. Nearly every eucaryotic cell type tested contained NMT, including avian, mammalian, insect, plant cells. Since NMT activity detected rabbit reticulocyte lysates, took advantage translational capability these determine precise point during translation at which myristate is attached pp60v-src. src mRNA, transcribed cloned v-src DNA, translated had been depleted endogenous myristate. Addition [3H]myristate 10 min after start synchronized resulted dramatic decrease incorporation radiolabeled into chains. These results imply that although can be posttranslationally substrates, myristylation vivo apparently very early cotranslational event occurs before 100 nascent chain are polymerized.