A novel cell adhesive protein engineered by insertion of the Arg-Gly-Asp-Ser tetrapeptide.
A tetrapeptide Arg-Gly-Asp-Ser (RGDS) has been shown to be a versatile cell recognition signal of extracellular matrix components for the interaction with cells. We introduced the RGDS tetrapeptide into a truncated form of protein A, a staphylococcal immunoglobulin-binding protein, by inserting an oligonucleotide cassette encoding the tetrapeptide into the coding region of the protein A expression vector pRIT2T. The mutagenized protein was capable of not only binding to immunoglobulin G but also mediating cell attachment and spreading onto an inert substrate. Cell adhesion mediated by the mutagenized protein was inhibitable by a synthetic peptide Gly-Arg-Gly-Asp-Ser but not by a related peptide Gly-Arg-Gly-Glu-Ser, confirming that the inserted RGDS tetrapeptide served as a recognition signal for cell adhesion. Furthermore, the RGDS-containing protein was capable of adhering cells onto an immunoglobulin-coated surface which could not by itself support cell adhesion. Thus, the cell adhesive and immunoglobulin binding activities of the mutagenized protein appear to function coordinately. The protocol described here is essentially applicable to any protein and, therefore, provides a general principle in tailoring novel multifunctional proteins having cell adhesive activity.
Maeda
T
T
Institute for Comprehensive Medical Science, Fujita-Gakuen Health University School of Medicine, Aichi, Japan.
Oyama
R
R
Ichihara-Tanaka
K
K
Kimizuka
F
F
Kato
I
I
Titani
K
K
Sekiguchi
K
K
D000595:Amino Acid Sequence; D000818:Animals; D000954:Antigens, Surface / Q000235:genetics*; D001483:Base Sequence; D002448:Cell Adhesion; D015815:Cell Adhesion Molecules; D002460:Cell Line; D004251:DNA Transposable Elements; D004926:Escherichia coli / Q000235:genetics; D005818:Genetic Engineering; D008969:Molecular Sequence Data; D009154:Mutation; D015345:Oligonucleotide Probes; D009842:Oligopeptides; D010957:Plasmids; D021382:Protein Sorting Signals / Q000235:genetics
D000954:Antigens, Surface; D015815:Cell Adhesion Molecules; D004251:DNA Transposable Elements; D015345:Oligonucleotide Probes; D009842:Oligopeptides; D021382:Protein Sorting Signals