Minimal FLAG sequence useful in the functional epitope tagging of H-Ras
Johnson, K.Y.; Liu, L.; Vincent, T.S.
Biotechniques 32(6): 1270 1272-6 1278 1280
2002
ISSN/ISBN: 0736-6205 PMID: 12074157 Document Number: 541841
Epitope tagging can interfere with normal protein function, indicating the need for an unobtrusive epitope tag. The FLAG(R) epitope (DYKDDDDK) was examined for a minimal epitope useful in the tagging of H-Ras. The heptapeptide tag, F7 (MDYKDDD), was found to retain reactivity with M2 and M5 monoclonal antibodies in immunoprecipitation, Western blotting, and immunofluorescence microscopy. The F7 tag did not interfere with Ras stability, EGF stimulation of Ras activation, and downstream phosphorylation of MAPK Erk1/2. Unlike the full FLAG sequence, the F7 tag had minimal effect on the growth properties of H-Ras in a colony-forming assay. The F7 tag may be useful when minimizing the effect of tagging on protein function is an important criterion in the selection of an N-terminal epitope tag.