NH2-terminal processing of actin in mouse L-cells in vivo.
When Dictyostellium discoideum actin is synthesized in vitro, it is made as a 43,000-dalton polypeptide with an NH2-terminal N-acetylmethionine. The acetylmethionine is then cleaved post-translationally, and the new NH2-terminal aspartic acid is acetylated to give the mature form of actin. Inhibition of methionine acetylation prevents methionine cleavage (Redman, K., and Rubenstein, P. (1981) J. Biol. Chem. 256, 13226-13229). In this paper, we describe the results of experiments designed to discover whether this novel actin processing pathway is peculiar to the rabbit reticulocyte lysate system or whether it is utilized by mammalian cells in vivo as well. We show that in mouse L-929 cells, actin is made as a 43,000-dalton protein with an NH2-terminal N-acylmethionine residue. Experiments using thin layer chromatography and digestion of the acylmethionine residue with hog kidney acylase I demonstrate that the acyl group is an acetyl residue. Pulse-chase experiments show that over the course of 1 h, this precursor is transformed first to an actin with a free NH2-terminal aspartic acid and is subsequently converted to mature L-cell actin with an acetylaspartic acid NH2 terminus. The half-life of the initial actin precursor in the cell appears to be approximately 12-15 min. These studies demonstrate the existence of this novel actin processing pathway in vivo and suggest that it is used for those actins where, in the gene, the initiator methionine codon directly precedes the codon for aspartic or glutamic acids, the residues normally found at the actin NH2 terminus.
D000199:Actins / Q000235:genetics* / Q000302:isolation & purification; D000595:Amino Acid Sequence; D000818:Animals; D004591:Electrophoresis, Polyacrylamide Gel; D007700:Kinetics; D007739:L Cells / Q000378:metabolism; D051379:Mice; D008970:Molecular Weight; D010446:Peptide Fragments / Q000032:analysis; D011499:Protein Processing, Post-Translational; D014357:Trypsin
D000199:Actins; D010446:Peptide Fragments; D014357:Trypsin