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Biomedical subjects

A Goetze

Publications and source records attributed to A Goetze.

10 recordsLinked to original sources
PubMed1995

HER4 expression correlates with cytotoxicity directed by a heregulin-toxin fusion protein.

We have constructed, expressed, and purified a fusion protein, HAR-TX beta 2, consisting of heregulin-beta 2 fused to a binding-defective form of Pseudomonas exotoxin A, PE40. The fusion protein was found to induce receptor tyrosine phosphorylation in CEM cells transfected with HER4 alone or in combination with HER2 but not in cells transfected with HER2 or HER1 alone. The phosphorylation of receptor tyrosines was both dose-dependent and saturable in amounts similar to those shown to be active for native heregulin. HAR-TX beta 2 was specifically cytotoxic toward a variety of carcinoma cell lines in the ng/ml range. However, some tumor cell lines were found to be insensitive to the cytotoxic action of the fusion protein even at > 2 micrograms/ml. Relative amounts of HER4, HER3, and HER2 were determined on seven cell lines sensitive and four cell lines insensitive to HAR-TX beta 2. All lines that express HER4 were killed by HAR-TX beta 2, while none lacking HER4 were affected. HAR-TX beta 2 was able to bind to and signal via tyrosine phosphorylation in cell lines that co-express HER2 and HER3 in the absence of HER4 without inducing cytotoxicity. Thus HAR-TX beta 2 may prove to be a useful reagent for the targeting and elimination of HER4-positive tumor cells.

ADP Ribose Transferases↗
PubMed1990

Discrimination of hepatitis B virus (HBV) subtypes using monoclonal antibodies to the PreS1 and PreS2 domains of the viral envelope.

We report the production and characterization of murine anti-PreS2 and anti-PreS1 monoclonal antibodies (mAb) and demonstrate their utility in discriminating hepatitis B virus (HBV) subtypes. On the basis of Western blotting and reciprocal competition binding to HBV virions, at least five distinct epitopes have been identified in the PreS domain: two within the PreS1 region and three within the PreS2 region. All PreS2 mAb bind M protein (gp33 and gp36) but only one group binds strongly to M and L proteins (p39 and gp42). This group determinant was mapped to peptide residues 120-145. The second group bound to an endoglycosidase F-sensitive epitope which is defined by a mannose-rich glycan at ASN 123 in the PreS2 region. The third group was mapped to peptide residues 150-174 and was reactive with the M envelope proteins but not L or S proteins on Western blots. All PreS1 mAb bind L protein but not M protein on Western blots. Using these mAb, HBV subtype assays were developed allowing evaluation of the Paris (1975) HBsAg subtype panel members along with other HBsAg-positive specimens. All Paris subtype members (except ayw2 and ayw3) could be easily distinguished by differential PreS2 mAb reactivity. The Paris subtypes, adw2, adw4, and adr, could be classified as distinct groups by PreS2 and PreS1 mAb binding. Specimens from Hong Kong and the United States classified as adw2 in the S region fell into two groups based on PreS2 mAb binding: one having reactivity similar to Paris adw2 subtype and the other having identical reactivity to Paris ayw1 subtype. Furthermore, some specimens classified as adr in the S region gave similar reactivity to the Paris ayr subtype in the PreS2 and PreS1 regions. One complicating factor in this approach toward subtyping was the discovery that some HBsAg positive sera may contain factors which block PreS epitopes. Grouping of HBV subtypes by PreS1, PreS2, and S mAb reactivity may allow better correlation with groupings based on HBV DNA sequence homology.

Animals↗
PubMed1989

Second generation assays for the detection of antibody to HBsAg using recombinant DNA-derived HBsAg.

A second generation radioimmunoassay (RIA) and enzyme-linked immunoassay (EIA) for the detection and quantitation of the antibody to hepatitis B surface antigen (anti-HBs) was developed which utilizes recombinant DNA-derived HBsAg (rHBsAg) in place of human plasma derived HBsAg. In these sandwich assays, rHBsAg immobilized on a solid phase was used to capture anti-HBs from the specimen and rHBsAg conjugated to horseradish peroxidase or radiolabeled with 125I was used as a detecting reagent. These rHBsAg-based assays were compared to a commercial radioimmunoassay for anti-HBs detection (AUSAB RIA). For a population of 1711 sera and plasma specimens, 99.2% overall agreement was demonstrated between the recombinant RIA and EIA and 98.6% agreement was observed between the recombinant assays and AUSAB-RIA. The recombinant assays demonstrated equivalent sensitivity and detectability to AUSAB RIA. Most discrepant samples were low-level reactive by AUSAB-RIA, generally less than 10 mIU/ml, and likely represent nonspecific reactivity since no other marker for hepatitis B infection was detected in these samples.

Autoimmune Diseases↗
PubMed1982

Isolation of platelet glycocalicin by affinity chromatography on thrombin-sepharose.

Platelet glycocalicin has been purified to homogeneity by a two step procedure involving affinity chromatography on WGA-Sepharose and then on thrombin-Sepharose using selective elution with heparin. The procedure is more rapid (3-4 days), more reproducible and gives about twice the yield (10 mg/40 units platelets) of the previous method (Okumura et al. J. Biol. Chem. 251, 5950-5955, 1976). The two preparations showed identical inhibition of aggregation of gel filtered platelets induced by thrombin and by ristocetin. Glycocalicin was cleaved in a controlled fashion by trypsin-Sepharose over 3hr to yield the macroglycopeptide and peptide "tail" fragments and there was no apparent further degradation with 18 hrs digestion.

Blood Platelets↗
PubMed1982

Structure of the high-affinity mast cell receptor for IgE.

Mast cells and related rat tumor basophils have a surface receptor that binds monomeric IgE with high avidity. The receptor in situ is unclustered, mobile, and univalent, and its aggregation into dimers and higher oligomers triggers degranulation. It has two subunits, alpha and beta. The alpha chain has a molecular weight of about 50,000, of which 30% is carbohydrate. Heterogeneity in the latter accounts at least in part and perhaps fully for the heterogeneity of the alpha chain. Studies with proteases have delineated two domains: alpha 1 appears slightly larger on sizing gels, and incorporation studies suggest it has more carbohydrate than alpha 2. The alpha 2 domain and a 24,000-dalton subfragment of it contain the principal site whose surface labeling is blocked by the presence of IgE. The alpha subunit is firmly but noncovalently bound to beta in a 1:1 complex in situ and in detergent extracts. Nevertheless, during thorough washing of IgE-receptor complexes with detergent, beta dissociates. A portion of beta, beta 1, is integrated in the membrane; it is this domain that is labeled when the receptor is isolated from cells reacted with the intramembranous probe iodonapthylnitrene and that interacts with the alpha chain. These results and others have been used to draw a provisional model of the receptor.

Animals↗
PubMed1981

Enzymatic cleavage products of the alpha subunit of the receptor for immunoglobulin E.

The high-affinity membrane receptor for immunoglobulin E on mast cells and on a tumor analogue, rat basophilic leukemia cells, consists of two polypeptide chains: an alpha chain of Mr congruent to 50,000 and a beta chain of Mr congruent to 30,000. In this study we reacted alpha chains purified from tumor cells with proteolytic and glycolytic enzymes and compared the products by using differential labeling procedures. A variety of proteolytic enzymes cleave the chain into two similar-sized fragments, alpha 1 and alpha 2. The alpha 1 fragment behaves as if it were slightly larger on electrophoresis through polyacrylamide gels and is rich in carbohydrate as determined by incorporation of [14C]glucosamine. Less incorporation is observed into alpha 2 but it, like alpha 1, binds to concanavalin A. Labeling of the surface proteins on intact cells by lactoperoxidase-catalyzed iodination leads to preferential, perhaps exclusive, labeling of the alpha 2 fragment. The relative proportion of incorporated 3H-labeled amino acids and radioactive Bolton--Hunter reagent suggests that the polypeptide portions of alpha 1 and alpha 2 are similar in size. From these and other data we propose that the alpha chain may be U-shaped. Results from endoglycosidase digestions show that the receptor as isolated is heterogeneous because of variable glycosylation.

Animals↗