A novel antibody against a synthetic NH2-terminal myristoyl glycyl src peptide can detect pp60v-src, the transforming protein of Rous sarcoma virus.
Novel antibodies were raised against a synthetic NH2-terminal myristoyl(Myr-) tetrapeptide(N-Myr-Gly-Ser-Ser-Lys) which is characteristic of an NH2-terminal portion of pp60v-src, the transforming protein of Rous sarcoma virus. Antisera raised against N-Myr-Gly-Ser-Ser-Lys-haemocyanin reacted with 125I-albumin conjugates with N-Myr-Gly-Ser-Ser-Lys. The immunoreaction was competed for by haemocyanin as well as albumin conjugated with this N-Myr-peptide, while underivatized proteins or an NH2-terminal octapeptide (Gly-Ser-Ser-Lys-Ser-Lys-Pro-Lys) had no effect. N-Myr-Gly-Ser-Ser-Lys-(125I)tyramine was also recognized by the antibody. The reaction was competed for by N-Myr-Gly-Ser-Ser-Lys, but not by Gly-Ser-Ser-Lys-Ser-Lys-Pro-Lys. These results suggest a high affinity of the antibody for an N-Myr-peptide moiety. The major (3H)myristate-labelled protein of an apparent molecular weight of 60,000 was detected from chick embryo fibroblasts transformed by Rous sarcoma virus (tsNY68). This protein was demonstrated to possess N-Myr-Gly-Ser-Ser-Lys by the immunoprecipitation and HPLC analyses. Furthermore, the entire circumference of the transformed cells was stained by the antibody upon an immunofluorescent microscopic observation. Thus, these results taken together indicate that the haptenic antibody raised against the myristoyl peptide is useful to detect pp60v-src protein.
Shoji
S
S
Department of Biochemistry, Faculty of Pharmaceutical Sciences, Kumamoto University, Japan.
Takenaka
O
O
Kida
Y
Y
Matsunaga
M
M
Hayashi
M
M
Horiuchi
S
S
Kubota
Y
Y
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000302:isolation & purification; D001358:Avian Sarcoma Viruses / Q000235:genetics*; D002478:Cells, Cultured; D002642:Chick Embryo; D002846:Chromatography, Affinity; D002851:Chromatography, High Pressure Liquid; D005455:Fluorescent Antibody Technique; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009842:Oligopeptides / Q000276:immunology / Q000302:isolation & purification; D015688:Oncogene Protein pp60(v-src) / Q000032:analysis* / Q000276:immunology; D011817:Rabbits / Q000276:immunology; D011863:Radioimmunoassay / Q000379:methods
D000906:Antibodies; D009842:Oligopeptides; D015688:Oncogene Protein pp60(v-src)