A synthetic approach to structure-function relationships in the murine epidermal growth factor molecule.

Heath, William F.; Merrifield, R. B.

doi:10.1073/pnas.83.17.6367

Cited by 85 publications

(30 citation statements)

References 23 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…The use of synthetic peptides to probe for functional domains of molecules has been applied by Heath and Merrifield (30), who have recently used solid-phase synthesis to identify the peptide sequences containing the active core of EGF. Similar studies with transforming growth factor type a have identified receptor binding domains (31).…”

Section: Resultsmentioning

confidence: 99%

Receptor- and heparin-binding domains of basic fibroblast growth factor.

Baird

Schubert

Ling

et al. 1988

Proc. Natl. Acad. Sci. U.S.A.

295

200

View full text Add to dashboard Cite

Two functional domains in the primary structure of basic fibroblast growth factor (FGF) have been identified on the basis of their ability to interact with the FGF receptor, bind radiolabeled heparin, and modulate the cellular response to FGF. Peptides derived from these two functional domains can act as partial agonists and antagonists in biological assays of FGF activity. Peptides related to the sequences of FGF-(24-68)-NH2 and FGF-(106-115)-NH2 inhibit thymidine incorporation into 3T3 fibroblasts when they are stimulated by FGF but have no effect when the cells are treated with either platelet-derived growth factor or epidermal growth factor. They also possess partial agonist activity and can stimulate DNA synthesis when tested in the absence of exogenous FGF. The active peptides have no effect on the binding of epidermal growth factor to its receptor on A431 cells and they can modulate the effects of FGF, but not fibronectin, on endothelial cell adhesion. The results suggest the possibility of designing specific analogs of FGF that are capable of inhibiting the biological effects of FGF.The molecular characterization of basic and acidic fibroblast growth factors (FGFs) has confirmed the existence of two classes of closely related angiogenic factors and helped establish the identity of an unusual family of mitogens (1-7). Acidic and basic FGFs exist in several molecular forms that are products of proteolytic processing at homologous sites (1)(2)(3)8). Both growth factors stimulate a wide spectrum of target cells derived from the primary and secondary mesenchyme as well as from the neural crest. The growth factors bind immobilized heparin with an unusually high affinity, a characteristic that has been exploited for their isolation and characterization from many tissues (4, 9). Because both FGFs also have the capacity to stimulate neovascularization (1-4), their physiological functions have been associated with reproduction, growth, and development. The possible identity of FGF-related proteins with the tumor angiogenic factor activities described by Folkman (10) has suggested that this family of mitogens may also play a critical role in several pathophysiological processes including the growth of tumors, diabetic proliferative retinopathies, and the woundhealing response. The ability of basic FGF to stimulate nerve regeneration (11) and to induce a neovascular response in the chicken chorioallantoic membrane (12), the rat brain (13), the kidney capsule (14), and the carotid artery (unpublished work) has suggested a possible therapeutic application of FGFs in stimulating tissue regeneration, the recovery from episodes of ischemia, and tissue transplantation.The identification of specific functional domains in the primary structure of basic FGF was examined as a first step in the design of recombinant analogs of basic FGF that have modified heparin-binding activity, increased biological activity, and/or antagonist activity. The results presented here identify two peptide sequences of interest (Fig. 1). M...

show abstract

Section: Resultsmentioning

confidence: 99%

Receptor- and heparin-binding domains of basic fibroblast growth factor.

Baird

Schubert

Ling

et al. 1988

Proc. Natl. Acad. Sci. U.S.A.

295

200

View full text Add to dashboard Cite

show abstract

“…Recombinant proteins, synthetic peptides, site-specific chemical derivatives, and proteolytic degradation have all been useful for generating altered molecules (14,31,42 AR precursor. Mature, secreted AR is synthesized as the middle portion of a 252-aa transmembrane precursor.…”

Section: Discussionmentioning

confidence: 99%

The amphiregulin gene encodes a novel epidermal growth factor-related protein with tumor-inhibitory activity.

Plowman¹,

Green²,

McDonald³

et al. 1990

Mol. Cell. Biol.

317

246

View full text Add to dashboard Cite

We have isolated the gene for a novel growth regulator, amphiregulin (AR), that is evolutionarily related to epidermal growth factor (EGF) and transforming growth factor a (TGF-a). AR is a bifunctional growth modulator: it interacts with the EGF/TGF-a receptor to promote the growth of normal epithelial cells and inhibits the growth of certain aggressive carcinoma cell lines. The 84-amino-acid mature protein is embedded within a 252-amino-acid transmembrane precursor, an organization similar to that of the TGF-a precursor. Human placenta and ovaries were found to express significant amounts of the 1.4-kilobase AR transcript, implicating AR in the regulation of normal cell growth. In addition, the AR gene was localized to chromosomal region 4q13-4q21, a common breakpoint for acute lymphoblastic leukemia.Cell growth and differentiation are regulated in part by the specific interaction of secreted growth factors and their membrane-bound receptors. Receptor-ligand interaction results in activation of intracellular signals leading to specific cellular responses. Epidermal growth factor (EGF), plateletderived growth factor, insulin, insulinlike growth factor 1, colony-stimulating factor 1, and fibroblast growth factor all transmit their growth-modulating signals by binding to and activating receptors with intrinsic tyrosine kinase activity (reviewed in reference 30). Characterization of the physiologic and chemical effects that result from the binding of EGF and transforming growth factor a (TGF-a) to the EGF receptor has served as a useful model for understanding receptor-ligand interactions, signal transduction, and the regulation of cell growth and oncogenesis (reviewed in reference 76).We have recently reported the purification and sequence analysis of a glycoprotein isolated from the conditioned media of 12-0-tetradecanoylphorbol-13-acetate (TPA)-treated MCF-7 cells (65,66). The protein was termed amphiregulin (AR) to reflect its bifunctional activities: it inhibits the growth of many human tumor cells, and it stimulates the proliferation of normal fibroblasts and keratinocytes. The secreted protein exists as a monomer of either 78 or 84 amino acids (aa), with the shorter form lacking the six N-terminal residues of the larger molecule. Sequence analysis reveals that AR has a region with striking homology to EGF (38%) and TGF-a (32%), yet it also has an N-terminal extension of 43 aa composed primarily of very basic, hydrophilic residues (Lys, Arg, and Asn). In addition, AR has functional homology with this class of growth factors; it partially competes for binding of EGF to the EGF receptor and can supplant the need for EGF or TGF-a to maintain keratinocytes in culture. AR differs from EGF and TGF-a in that it fails to promote anchorage-independent growth of normal rat kidney (NRK) fibroblasts in the presence of TGF-P and inhibits the growth of certain tumor cells that proliferate in response to EGF or TGF-a.In this report, we describe the isolation and characterization of cDNA and genomic clones for human AR, t...

show abstract

“…Both Meb and Acm were used for blocking Cys. To remove the peptide from the resin, a low/high H F method (Tam et al, 1983) (Heath & Merrifield, 1986). The peptide was purified to homogeneity by preparative HPLC.…”

Section: Peptide Synthesismentioning

confidence: 99%

Two‐step selective formation of three disulfide bridges in the synthesis of the C‐terminal epidermal growth factor‐like domain in human blood coagulation factor IX

et al. 1994

View full text Add to dashboard Cite

The 45-residue C-terminal EGF-like domain in human blood coagulation factor IX has been synthesized by a 2-step method to form selectively 3 disulfide bridges. Four out of 6 cysteines are blocked with either trityl or 4-methylbenzyl, and the remaining 2 cysteines are blocked with acetamidomethyl (Acm). In the first step, 4 free cysteinyl thiols are released concurrently with the removal of all protecting groups except Acm and are oxidized to form 1 of the 3 possible isomers containing 2 pairs of disulfides. In the second step, iodine is used to remove the Acm groups to yield the third disulfide bridge. This approach reduces the number of possible disulfide bridging patterns from 15 to 3. To determine the optimal protecting group strategy, 3 peptides are synthesized, each with Acm blocking

show abstract

A synthetic approach to structure-function relationships in the murine epidermal growth factor molecule.

Cited by 85 publications

References 23 publications

Receptor- and heparin-binding domains of basic fibroblast growth factor.

Receptor- and heparin-binding domains of basic fibroblast growth factor.

The amphiregulin gene encodes a novel epidermal growth factor-related protein with tumor-inhibitory activity.

Two‐step selective formation of three disulfide bridges in the synthesis of the C‐terminal epidermal growth factor‐like domain in human blood coagulation factor IX

Contact Info

Product

Resources

About