The N-terminal SH4 region of the Src family kinase Fyn is modified by methylation and heterogeneous fatty acylation: Role in membrane targeting, cell adhesion, and spreading Journal Article
| Authors: | Liang, X.; Lu, Y.; Wilkes, M.; Neubert, T. A.; Resh, M. D. |
| Article Title: | The N-terminal SH4 region of the Src family kinase Fyn is modified by methylation and heterogeneous fatty acylation: Role in membrane targeting, cell adhesion, and spreading |
| Abstract: | The N-terminal SH4 domain of Src family kinases is responsible for promoting membrane binding and plasma membrane targeting. Most Src family kinases contain an N-terminal Met-Gly-Cys consensus sequence that undergoes dual acylation with myristate and palmitate after removal of methionine. Previous studies of Src family kinase fatty acylation have relied on radiolabeling of cells with radioactive fatty acids. Although this method is useful for verifying that a given fatty acid is attached to a protein, it does not reveal whether other fatty acids or other modifying groups are attached to the protein. Here we use matrix-assisted laser desorption/ ionization-time of flight (MALDI-TOF) mass spectrometry to identify fatty acylated species of the Src family kinase Fyn. Our results reveal that Fyn is efficiently myristoylated and that some of the myristoylated proteins are also heterogeneously S-acylated with palmitate, palmitoleate, stearate, or oleate. Furthermore, we show for the first time that Fyn is trimethylated at lysine residues 7 and/or 9 within its N-terminal region. Both myristoylation and palmitoylation were required for methylation of Fyn. However, a general methylation inhibitor had no inhibitory effect on myristoylation and palmitoylation of Fyn, suggesting that methylation occurs after myristoylation and palmitoylation. Lysine mutants of Fyn that could not be methylated failed to promote cell adhesion and spreading, suggesting that methylation is important for Fyn function. |
| Keywords: | methylation; proto-oncogene proteins; nonhuman; mass spectrometry; animal cell; animals; gene targeting; protein tyrosine kinase; structure-activity relationship; cercopithecus aethiops; cos cells; animalia; luminescent proteins; amino acid sequence; amino terminal sequence; recombinant fusion proteins; isotope labeling; peptide fragments; cell membrane; green fluorescent proteins; cell fractionation; fatty acids; matrix assisted laser desorption ionization time of flight mass spectrometry; biochemistry; cell adhesion; mutagenesis; enzymes; lysine; rna transport; enzyme modification; protein modification; membrane binding; spectrometry, mass, matrix-assisted laser desorption-ionization; acylation; palmitoylation; consensus sequence; protein kinase fyn; proto-oncogene proteins c-fyn; myristic acid; myristylation; oleic acid; ionization; biological membranes; desorption; acyltransferases; palmitic acid; species differentiation; cell spreading; plasma membrane; stearic acid; priority journal; article; laser beam effects; palmitoleic acid |
| Journal Title: | Journal of Biological Chemistry |
| Volume: | 279 |
| Issue: | 9 |
| ISSN: | 0021-9258 |
| Publisher: | American Society for Biochemistry and Molecular Biology |
| Date Published: | 2004-02-27 |
| Start Page: | 8133 |
| End Page: | 8139 |
| Language: | English |
| DOI: | 10.1074/jbc.M311180200 |
| PROVIDER: | scopus |
| PUBMED: | 14660555 |
| DOI/URL: | |
| Notes: | J. Biol. Chem. -- Cited By (since 1996):43 -- Export Date: 16 June 2014 -- CODEN: JBCHA -- Source: Scopus |
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