Characterization of sequences within heparin-binding EGF-like growth factor that mediate interaction with heparin.
Heparin-binding (HB) epidermal growth factor (EGF)-like growth factor (HB-EGF), a member of the EGF protein family, is a potent mitogen for fibroblasts, smooth muscle cells, and keratinocytes that was initially identified as a secreted product of macrophage-like cells. HB-EGF and EGF appear to act on target cells utilizing the same receptor, but HB-EGF is distinguishable from EGF by its strong affinity for heparin. To facilitate studies of structure-function relationships in HB-EGF, a bacterial recombinant expression system was established that produced biologically active HB-EGF with the expected disulfide bonding pattern. Mutagenesis and protease digestion studies of the recombinant HB-EGF, coupled with heparin-binding analyses of synthetic peptides, indicated that the sequences within HB-EGF mediating its interaction with heparin are located primarily in a stretch of 21 amino acids characterized by a high content of lysine and arginine residues. Most of this heparin-binding domain lies in an amino-terminal region of HB-EGF that has no counterpart in EGF, but a portion of the 21-residue sequence extends into the EGF-like region of HB-EGF. In addition, the mutagenesis and synthetic peptide studies indicated that sequences in HB-EGF lying outside of the 21-residue stretch can also influence the interaction with heparin. Finally, a synthetic peptide derived from the 21-residue stretch was found to compete with HB-EGF for binding to Chinese hamster ovary cells, suggesting that the heparin-binding sequences in HB-EGF may also mediate the interaction of this factor with cell surface heparan sulfate proteoglycan.
Thompson
S A
SA
Scios Nova Inc., Mountain View, California 94043.
Higashiyama
S
S
Wood
K
K
Pollitt
N S
NS
Damm
D
D
McEnroe
G
G
Garrick
B
B
Ashton
N
N
Lau
K
K
Hancock
N
N
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D016466:CHO Cells; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D006224:Cricetinae; D004815:Epidermal Growth Factor / Q000302:isolation & purification / Q000378:metabolism* / Q000494:pharmacology*; D004926:Escherichia coli; D006493:Heparin / Q000378:metabolism*; D066257:Heparin-binding EGF-like Growth Factor; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins; D051379:Mice; D008934:Mitogens / Q000494:pharmacology*; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism / Q000494:pharmacology; D014407:Tumor Cells, Cultured
C587883:HBEGF protein, human; C587884:Hbegf protein, mouse; D066257:Heparin-binding EGF-like Growth Factor; D036341:Intercellular Signaling Peptides and Proteins; D008934:Mitogens; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D004815:Epidermal Growth Factor; D006493:Heparin