Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P04626 (erbB-2)
 5,251 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In this work, we have used Xenopus oocyte maturation as a read-out for examining the ability of the neu tyrosine kinase (p185neu) to participate with the epidermal growth factor (EGF) receptor in a common signal transduction pathway. We find that unlike the case for the EGF receptor, which elicits EGF-dependent maturation of these oocytes as reflected by their germinal vesicle breakdown (GVBD), neither the normal neu tyrosine kinase (p185val664) nor the oncogenic form of neu (p185glu664) are able to effectively trigger this maturation event. However, expression of p185glu664 causes a specific and significant promotion of the progesterone-induced GVBD, reducing the half-time for this maturation even from approximately 9 h to approximately 5 h. Stimulation of the progesterone-induced GVBD did not occur following the expression of a kinase-deficient p185neu protein (in which a lysine residue at position 758 was changed to alanine). Essentially identical results were obtained when the mRNAs coding for fusion proteins comprised of the extracellular domain of the receptor for immunoglobulin E (IgE), and the membrane-spanning and tyrosine kinase domains of normal or oncogenic p185neu (designated IgER/p185val664 and IgER/p185glu664, respectively), were injected into oocytes. Antigen-induced crosslinking of IgER/p185val164 proteins expressed in oocytes caused a reduction in the half-time for the progesterone-stimulated GVBD from approximately 9 h to approximately 7 h. Thus, the aggregation of the membrane-spanning and/or tyrosine kinase domains of p185val664 partially mimics the effects of the oncogenic forms of p185neu. Overall, the results of these studies suggest that the activation of the p185neu tyrosine kinase by a point mutation within its membrane-spanning helix, or an aggregation event, can result in the facilitation of oocyte maturation events that are elicited by other factors (e.g. progesterone). However, the activated p185neu tyrosine kinases are not able to mimic the EGF-stimulated EGF receptor tyrosine kinase in triggering oocyte maturation, which suggests that the EGF receptor and the p185neu tyrosine kinase do not input into identical signal transduction pathways in these cells.
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PMID:The effects of the normal and oncogenic forms of the neu tyrosine kinase, and the corresponding forms of an immunoglobulin E receptor/neu tyrosine kinase fusion protein, on Xenopus oocyte maturation. 135 69

Four residues in the carboxy-terminal domain of human epidermal growth factor (hEGF), glutamate 40, glutamine 43, arginine 45, and aspartate 46 were targeted for site-directed mutagenesis to evaluate their potential role in epidermal growth factor (EGF) receptor-ligand interaction. One or more mutations were generated at each of these sites and the altered recombinant hEGF gene products were purified and evaluated by radioreceptor competition binding assay. Charge-conservative replacement of glutamate 40 with aspartate resulted in a decrease in receptor binding affinity to 30% relative to wild-type hEGF. On the other hand, removal of the electrostatic charge by substitution of glutamate 40 with glutamine or alanine resulted in only a slightly greater decrease in receptor binding to 25% relative receptor affinity. The introduction of a positive charge upon substitution of glutamine 43 with lysine had no effect on receptor binding. The substitution of arginine 45 with lysine also showed no effect on receptor binding, unlike the absolute requirement observed for the arginine side-chain at position 41 [Engler DA, Campion SR, Hauser MR, Cook JS, Niyogi, SK: J Biol Chem 267:2274-2281, 1992]. Subsequent elimination of the positive charge of lysine 45 by reaction with potassium cyanate showed that the electrostatic property of the residue at this site, as well as that at lysine 28 and lysine 48, was not required for receptor-ligand association.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Evaluation of the role of electrostatic residues in human epidermal growth factor by site-directed mutagenesis and chemical modification. 135 99

Previous studies have shown that lysine- and arginine-rich proteins can enhance the activity of tyrosine and serine/threonine protein kinases. However, the kinetics and mechanism of this activation are not fully understood. Therefore we investigated the ability of poly(amino acids) and the arginine-rich protein, protamine, to alter the kinetic properties of epidermal growth factor (EGF) receptor protein-tyrosine kinase activity using immunoaffinity-purified receptor isolated from human epidermoid carcinoma (A431) cells. Poly(L-lysine), poly(L-arginine) and protamine stimulated EGF receptor kinase activity by 3-5-fold at non-saturating doses of ATP and peptide substrate, while poly(L-glutamate) had no effect. Initial kinetic studies demonstrated an increase in the maximum velocity and a decrease in the apparent Km for the peptide substrate angiotensin II in the presence of the basic effectors. Further analysis of the kinetic mechanism by product inhibition revealed that protamine altered the pattern of ADP inhibition towards the peptide substrate but not towards ATP. The change was indicative of the receptor's ability to form an enzyme-angiotensin II-ADP ternary complex in the presence of protamine but not in its absence. In addition, the basic effectors had a substantially decreased influence on the kinase activity of a C-terminally truncated form of the EGF receptor. Thus the changes in kinase activity may be partially mediated by the C-terminal region of the receptor, which contains the sites of receptor self-phosphorylation. These results suggest that the basic domains of proteins can interact with the EGF receptor to induce changes in its kinetic properties, especially with regard to reactant recognition and binding.
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PMID:Alteration of the kinetic properties of the epidermal growth factor receptor tyrosine kinase by basic proteins. 137 Jun 7

In a preliminary study we demonstrated that the formation of the epidermal growth factor (EGF) receptor-ligand complex requires the participation of the highly conserved arginine 41 side chain of the growth factor peptide (Engler, D.A., Montelione, G.T., and Niyogi, S.K. (1990) FEBS Lett. 271, 47-50). In an attempt to gain further insight into the nature of this interaction(s), we used both site-directed mutagenesis and chemical modification reagents to produce human EGF (hEGF) analogues with altered chemical properties of the residue 41 side chain. Eight mutant analogues of hEGF were generated, substituting arginine 41 with lysine, glutamine, isoleucine, tyrosine, glycine, alanine, aspartate, or glutamate. Although each of the mutant analogues was able to displace wild-type hEGF fully in receptor competition binding assays, affinity of the receptor for the mutants was substantially reduced, varying from 0.4 to less than 0.01% of that observed for wild-type growth factor. At sufficiently high concentrations these mutants were able to stimulate DNA synthesis in mouse keratinocytes. Substitution of lysine for arginine 41 reduced the receptor affinity 250-fold from that observed for wild type, despite retention of the positive electrostatic charge. The lysine substitution leaves a reactive amine at position 41 and made it possible, using amine-specific chemical modification reagents, to produce selected arginine homologues that were tested for their effects on receptor binding, receptor tyrosine kinase activation, and stimulation of DNA synthesis in mouse keratinocytes. The reaction of lysine 41 with methyl acetimidate resulted in a lysineacetamidine product which only partially restored activity of the lysine hEGF mutant. However, reaction with O-methylisourea resulted in generation of an arginine 41 homologue (homoarginine) which restored full activity. The results indicate that the chemical properties inherent in the guanidinium group of the arginine 41 side chain of hEGF are responsible for optimal receptor-ligand association.
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PMID:Critical functional requirement for the guanidinium group of the arginine 41 side chain of human epidermal growth factor as revealed by mutagenic inactivation and chemical reactivation. 173 35

Transforming growth factor-alpha (TGF-alpha) is a growth-promoting protein that binds to the epidermal growth factor (EGF) receptor. To identify critical residues that govern TGF-alpha-EGF receptor binding, we prepared site-specific substitution mutants of TGF-alpha. Mutant proteins were tested in receptor-binding and mitogenesis assays. Semiconservative substitutions at positions 4, 12, 18, and 45 decreased biological activity 2.1- to 14-fold. The conservative substitution of lysine for arginine at position 42 completely eliminated biological activity. Amino acid composition analysis of proteolytic fragments from TGF-alpha and the Lys-42 mutant indicated that these proteins contained the same disulfide bonds. These studies suggest that arginine 42 may be a contact point for TGF-alpha-EGF receptor interaction.
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PMID:Substitution of lysine for arginine at position 42 of human transforming growth factor-alpha eliminates biological activity without changing internal disulfide bonds. 250 41

A variant human epidermal growth factor (EGF) receptor (HER) with a transition (G to A) at codon 497, resulting in a substitution of a lysine for an arginine, was recently demonstrated in several human pancreatic cancer cell lines. In the present study, we compared the frequency of expression of wild-type (HER497R) and variant (HER497K) EGF receptors in normal and malignant pancreatic tissue samples using single-strand conformation polymorphism analysis. Both receptors were expressed in normal (42%) and malignant (46%) tissues. Three samples (two normal and one cancer) expressed only HER497K. The cancer sample that was homozygous for HER497K exhibited strong EGF receptor immunoreactivity. The high frequency of HER497K expression suggests that it is due to a relatively common polymorphism in the HER coding region. Its presence in some of the cancer samples suggests that, like the wild-type receptor, HER497K also has a role in pancreatic cancer.
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PMID:Single-strand conformation polymorphism analysis of the epidermal growth factor receptor at codon 497. 883 Mar 26

Detergent-permeabilized EGFR-T17 fibroblasts, which overexpress the human epidermal growth factor (EGF) receptor, phosphorylate both poly-L-(glutamic acid, tyrosine) and exogenous calmodulin in an EGF-stimulated manner. Phosphorylation of calmodulin requires the presence of cationic polypeptides, such as poly-L-(lysine) or histones, which exert a biphasic effect toward calmodulin phosphorylation. Optimum cationic polypeptide/calmodulin molar ratios of 0.3 and 7 were determined for poly-L-(lysine) and histones, respectively. Maximum levels of calmodulin phosphorylation were attained in the absence of free calcium, and a strong inhibition of this process was observed at very low concentrations (Ki = 0.2 microM) of this cation. The incorporation of phosphate into calmodulin occurred predominantly on tyrosine residue(s) and was stimulated 34-fold by EGF.
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PMID:Phosphorylation of calmodulin by permeabilized fibroblasts overexpressing the human epidermal growth factor receptor. 904 62

The adaptor protein Shc contains a phosphotyrosine binding (PTB) domain and a Src homology 2 (SH2) domain, both of which are known to interact with phosphorylated tyrosines. We have shown previously that tyrosine 1148 of the activated epidermal growth factor (EGF) receptor is a major binding site for Shc while tyrosine 1173 is a secondary binding site in intact cells. In the present study, we investigated the interaction between the PTB and SH2 domains of Shc and the activated human EGF receptor. Mutant 52-kDa Shc with an arginine-to-lysine substitution at residue 175 in the PTB domain (Shc R175K) or 397 in the SH2 domain (Shc R397K) was coexpressed in Chinese hamster ovary cells overexpressing the wild-type or mutant EGF receptors that retained only one of the autophosphorylation sites at tyrosine 1148 (QM1148) or 1173 (QM1173). Shc R397K was coprecipitated with the QM1148 and QM1173 receptors, was tyrosine-phosphorylated, and associated with Grb2 and Sos. In contrast, coprecipitation of Shc R175K with the mutant receptors was barely detectable. In cells expressing the QM1173 receptor, Shc R175K was tyrosine-phosphorylated and associated with Grb2, while association of Sos was barely detectable. In cells expressing the QM1148 receptor, tyrosine phosphorylation of Shc R175K was markedly reduced. When both Shc R175K and 46-kDa Shc R397K were coexpressed with the mutant receptors, p46 Shc R397K was dominantly tyrosine-phosphorylated. In cells expressing the wild-type receptor, Shc R397K, but not Shc R175K, translocated to the membrane in an EGF-dependent manner. In addition, Ras activity stimulated by the immunoprecipitates of Shc R397K was significantly higher than that by the immunoprecipitates of Shc R175K. The present results indicate that tyrosine 1148 of the activated EGF receptor mainly interacts with the Shc PTB domain in intact cells. Tyrosine 1173 interacts with both the PTB and SH2 domains, although the interaction with the PTB domain is dominant. In addition, Shc bound to the activated EGF receptor via the PTB domain dominantly interacts with Grb2-Sos complex and plays a major role in the Ras-signaling pathway.
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PMID:Shc phosphotyrosine-binding domain dominantly interacts with epidermal growth factor receptors and mediates Ras activation in intact cells. 954 89

The "Fab immunogene" is a novel gene transfer vehicle in which the Fab fragment of anti-human epidermal growth factor (EGF) receptor antibody B4G7 is conjugated with poly-L-lysine to form an affinity complex with DNA. It was developed to target delivery of therapeutic genes into EGF receptor-hyperproducing tumor cells. Various characteristic features of the immunogene have been documented (Chen et al., 1998). Here we add further evidence to prove that in vitro transfer of beta-galactosidase/Fab immunogene is exclusively to EGF receptor-positive cells and that the herpes simplex virus thymidine kinase (TK)/Fab immunogene induces substantial suicide effects on A431 tumor cells when treated together with ganciclovir. The in vivo specificity of the immunogene transfer was examined using A431 tumor-bearing nude mice. When these nude mice were injected intraperitoneally with the chloramphenicol acetyltransferase (CAT)/Fab immunogene, CAT DNA was detected in the tumors as well as in liver and kidney but not brain, whereas CAT mRNA and enzyme activity were detected only in the tumors. Local and intraperitoneal injection of the TK/Fab immunogene and subsequent administration of ganciclovir effectively suppressed the growth of A431 tumors transplanted on the backs of nude mice. These observations suggest a possible application of the Fab immunogene system in cancer gene therapy.
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PMID:Targeted in vivo delivery of therapeutic gene into experimental squamous cell carcinomas using anti-epidermal growth factor receptor antibody: immunogene approach. 987 65

The Fab fragment of monoclonal antibody B4G7 against human epidermal growth factor (EGF) receptor was conjugated with cationic poly-L-lysine and the resulting conjugate was further complexed with reporter genes or therapeutic genes. This Fab/DNA complex was designated as "Fab immunogene." The Fab immunogene transfer in vitro was mediated through the EGF receptors in two melanoma cell lines. The frequency of cells expressing beta-galactosidase (beta-Gal) reporter gene was approximately 1%. The induction of suicide effects after Fab immunogene transfer of herpes simplex virus thymidine kinase (TK) or Escherichia coli cytosine deaminase (CD) gene was quite remarkable, and the growth of melanoma cells was inhibited for over 7 days in the presence of ganciclovir (GCV) or 5-fluorocytosine (5-FC). Similarly, when melanoma cells treated in vitro with the Fab immunogene carrying TK or CD were transplanted into the back of nude mouse, subsequent systemic administration of GCV or 5-FC effectively suppressed the growth of tumors, indicating the occurrence of in vivo suicide effects.
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PMID:Ex vivo delivery of suicide genes into melanoma cells using epidermal growth factor receptor-specific Fab immunogene. 1036 86


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