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M S Co

Publications and source records attributed to M S Co.

29 records · Page 2Linked to original sources
PubMed1988

Distinct binding sites for zinc and double-stranded RNA in the reovirus outer capsid protein sigma 3.

By atomic absorption analysis, we determined that the reovirus outer capsid protein sigma 3, which binds double-stranded RNA (dsRNA), is a zinc metalloprotein. Using Northwestern blots and a novel zinc blotting technique, we localized the zinc- and dsRNA-binding activities of sigma 3 to distinct V8 protease-generated fragments. Zinc-binding activity was contained within an amino-terminal fragment that contained a transcription factor IIIA-like zinc-binding sequence, and dsRNA-binding activity was associated with a carboxy-terminal fragment. By these techniques, new zinc- and dsRNA-binding activities were also detected in reovirus core proteins. A sequence similarity was observed between the catalytic site of the picornavirus proteases and the transcription factor IIIA-like zinc-binding site within sigma 3. We suggest that the zinc- and dsRNA-binding activities of sigma 3 may be important for its proposed regulatory effects on viral and host cell transcription and translation.

Amino Acid Sequence↗
PubMed1986

Nucleic acid sequence of an internal image-bearing monoclonal anti-idiotype and its comparison to the sequence of the external antigen.

The monoclonal anti-idiotypic antibody (mAb2) 87.92.6 directed against the 9B.G5 antibody specific for the virus neutralizing epitope on the mammalian reovirus type 3 hemagglutinin was previously demonstrated to express an internal image of the receptor binding epitope of the reovirus type 3. Furthermore, this mAb2 has autoimmune reactivity to the cell surface receptor of the reovirus. The nucleotide and deduced amino acid sequences of the 87.92.6 mAb2 heavy and light chains are described in this report. The sequence analysis reveals that the same heavy chain variable and joining (VH and JH) gene segments are used by the 87.92.6 anti-idiotypic mAb2 and by the dominant idiotypes of the BALB/c anti-GAT (cGAT) and anti-NP (NPa) responses. [GAT; random polymer that is 60% glutamic acid, 30% alanine, and 10% tyrosine. NP; (4-hydroxy-3-nitrophenyl)-acetyl.] Despite extensive homology at the level of the heavy chain variable regions, the NPa positive BALB/c anti-NP monoclonal antibody 17.2.25 binds neither 9B.G5 nor the cellular receptor for the hemagglutinin. Amino acid sequence comparison between the viral hemagglutinin and the 87.92.6 mAb2 light chain "internal image," reveals an area of significant homology indicating that antigen mimicry by antibodies may be achieved by sharing primary structure.

Amino Acid Sequence↗
PubMed1985

Anti-idiotypic antibody identifies the cellular receptor of reovirus type 3.

The binding and subsequent infectivity of reovirus to target cells are mediated by interaction with specific cell surface viral receptors. To gain a more detailed understanding of the biochemistry of the reovirus receptor and the cellular consequences of viral attachment, we have studied the binding of type 3 reovirus (Dearing strain) in a quantitative manner utilizing an antiidiotypic antibody probe. A syngeneic monoclonal antiidiotypic antibody (87.92.6) was prepared by immunization with hybridoma cells which secrete an antireovirus hemagglutinin-specific antibody. This antiidiotypic antibody was previously shown to specifically recognize the cell surface receptor for reovirus type 3. In this report, we demonstrate that antiidiotype mimicked reovirus tropism in binding to murine thymomas; antiidiotype inhibited the binding of reovirus to specific targets, but not the binding of anti-H-2; and cross linking of receptor-bound antiidiotype by antiimmunoglobulin induced patching, but not capping of reovirus receptors. Utilizing radiolabeled antiidiotype, we next quantitate the number of reovirus receptors on R1.1 and YAC thymoma cells and, finally, report on the preliminary identification of the reovirus receptor as a 67,000-Da membrane glycoprotein.

Animals↗
PubMed1985

Structural similarities between the mammalian beta-adrenergic and reovirus type 3 receptors.

The mechanism by which viruses bind to and infect specific tissues to cause disease has only recently begun to be understood. The mammalian reoviruses provide an especially attractive model for studying the details of cell surface recognition. The cell and tissue tropism of reovirus is determined by a portion of the viral hemagglutinin termed the neutralization domain. We have reported previously on the generation of both monoclonal and polyclonal anti-idiotypic antibodies that mimic the viral hemagglutinin in the specificity of binding to the reovirus receptor. By using these anti-idiotypic antibodies as specific probes, we have successfully isolated the mammalian reovirus receptor from neuronal and lymphoid cells. In the present study, we report that the reovirus receptor is structurally similar to the mammalian beta-adrenergic receptor. This conclusion is based on the following observations: (i) purified beta-adrenergic receptor is immunoprecipitable by anti-reovirus receptor antibody; (ii) purified reovirus receptor obtained from murine thymoma cells and beta-adrenergic receptor obtained from calf lung exhibit identical molecular masses and isoelectric points; (iii) trypsin digests of purified reovirus and beta-adrenergic receptors display indistinguishable fragment patterns; (iv) purified reovirus receptor binds the beta-antagonist [125I]iodohydroxybenzylpindolol and this binding is blocked by the beta-agonist isoproterenol.

Animals↗
PubMed1985

Isolation and biochemical characterization of the mammalian reovirus type 3 cell-surface receptor.

A cell-surface receptor for the mammalian reovirus type 3 hemagglutinin was isolated by using antiidiotypic anti-receptor antibodies. The receptor is a glycoprotein with a molecular mass of 67,000 daltons and a pI of 5.9. Evidence that the isolated structure represents the reovirus receptor was obtained by electrophoretic immunoblot studies, which demonstrated that the 67,000-dalton glycoprotein is the only cell-surface structure recognized by both reovirus type 3 and the anti-receptor immunoglobulin. Comparison of the reovirus receptor on murine thymoma (R1.1) and rat neuroblastoma (B104) cells indicated that similar structures on the cell surface are recognized by the reovirus type 3 and the anti-receptor antibodies as previously suggested from cellular and binding studies. This receptor was found on mouse, rat, monkey, and human cells. Furthermore, diverse tissue types, including lymphoid and neuronal cells, express the receptor structure. The receptor structure is discussed in terms of its role in mediating viral tropism and as an essential cell-surface protein.

Animals↗
PubMed1984

Anti-idiotypic antibodies as probes of cell surface receptors.

Anti-idiotypic antibodies have proven to have unique applications as probes in both functional and biochemical studies of cell surface receptors. Anti-idiotypic receptor antibodies have been prepared to antibodies which bind to purified ligand, as in the case of insulin, retinol-binding protein, the mammalian reovirus receptor, and the neutrophil chemotactic receptor, and to natural ligand analogs, such as the beta-adrenergic antagonist alprenolol. These systems have documented the usefulness of anti-idiotypic antibodies in the quantitation and modulation of specific membrane receptors on a variety of cell types. Anti-idiotypic antibodies have also been utilized for the isolation of specific membrane receptors, e.g., reovirus and B-1H globulin receptors. Some anti-idiotypic receptor antibodies, e.g., insulin and reovirus systems, have been shown to mimic the physiological properties of ligand upon binding to cellular receptors. These antibodies enable a new dimension of both receptor based cellular studies and therapeutic regimens. This review focuses on the past use of anti-idiotypic antibodies as probes of cell surface receptors, and on the methodologies required for the successful application of anti-idiotypic antibodies for use in further membrane receptor studies, and of the genes which encode and regulate these receptors. We also discuss the use of anti-idiotypic antibodies in the understanding of and therapeutic approach to receptor related diseases.

Animals↗
PubMed1983

Quantitative Cu(I) determination using X-ray absorption edge spectroscopy: oxidation of the reduced binuclear copper site in type 2 depleted Rhus laccase.

We report a procedure, through difference comparison of X-ray absorption edge spectra, for the quantitative determination of Cu(I) content in copper complexes of mixed oxidation state composition. This technique is tested on copper model systems and then used to quantitatively determine that untreated T2D Rhus laccase contains 70 +/- 15% Cu(I). Whereas excess ferricyanide is demonstrated not to alter the Cu(I) content of the untreated T2D, aqueous peroxide and nitrite at pH 6.0 are shown to oxidize the cuprous type 3 site and generate met T2D protein forms.

Binding Sites↗
PubMed1983

EXAFS investigation of the binuclear cupric site in met T2D Rhus laccase and its azide bound derivative.

EXAFS analysis of met T2D Rhus laccase and its azide bound derivative indicates an average of 0.33 S at 2.09 A and 3-4 N (or O) atoms at 2.00 A per copper atom for the three copper centers. Using the plastocyanin Cu(II) EXAFS spectrum to model the type 1 site in laccase, a difference EXAFS spectrum for the type 3 site is generated; this spectrum enables assignment of the one S ligand in met T2D to the type 1 site and indicates no evidence of a detectable copper scatterer for the coupled binuclear copper site. Implications regarding type 3 optical features and related studies on the hemocyanins are also discussed.

Azides↗
PubMed1982

X-ray absorption spectroscopy of the dimeric iron site in azidomethemerythrin from Phascolopsis gouldii.

X-ray absorption spectroscopy has been used to study the dimeric iron center in azidomethemerythrin from Phascolopsis gouldii. Absorption edge data confirm that the two iron atoms are present as Fe(III) and suggest a hexa-coordination site for each of the iron atoms. The extended x-ray absorption fine structure (EXAFS) analysis provides direct structural evidence of a mu-oxo bridge between the two iron atoms at an average Fe-O distance of 1.71-1.76 A. Analysis using a multiple-scattering formalism calculates upper limits of 165 degrees for the Fe-O-Fe bridging angle and 3.38 A for the Fe-Fe distance. This result agrees with current crystallographic models being determined by refinement of structures of two azidomethemerythrins.

Animals↗
PubMedDate not supplied

Comparison of the three-dimensional structures of a humanized and a chimeric Fab of an anti-gamma-interferon antibody.

The objective of this work is to compare the three-dimensional structures of "humanized" and mouse-human chimeric forms of a murine monoclonal antibody elicited against human gamma-interferon. It is also to provide structural explanations for the small differences in the affinities and biological interactions of the two molecules for this antigen. Antigen-binding fragments (Fabs) were produced by papain hydrolysis of the antibodies and crystallized with polyethylene glycol (PEG) 8,000 by nearly identical microseeding procedures. Their structures were solved by X-ray analyses at 2.9 A resolution, using molecular replacement methods and crystallographic refinement. Comparison of these structures revealed marked similarities in the light (L) chains and near identities of the constant (C) domains of the heavy (H) chains. However, the variable (V) domains of the heavy chains exhibited substantial differences in the conformations of all three complementarity-determining regions (CDRs), and in their first framework segments (FR1). In FR1 of the humanized VH, the substitution of serine for proline in position 7 allowed the N-terminal segment (designated strand 4-1) to be closely juxtaposed to an adjacent strand (4-2) and form hydrogen bonds typical of an antiparallel beta-pleated sheet. The tightening of the humanized structure was relayed in such a way as to decrease the space available for the last portion of HFR1 and the first part of HCDR1. This compression led to the formation of an alpha-helix involving residues 25-32. With fewer steric constraints, the corresponding segment in the chimeric Fab lengthened by at least 1 A to a random coil which terminated in a single turn of 310 helix. In the humanized Fab, HCDR1, which is sandwiched between HCDR2 and HCDR3, significantly influenced the structures of both regions. HCDR2 was forced into a bent and twisted orientation different from that in the chimeric Fab, both at the crown of the loop (around proline H52a) and at its base. As in HCDR1, the last few residues of HCDR2 in the humanized Fab were compressed into a space-saving alpha-helix, contrasting with a more extended 310 helix in the chimeric form. HCDR3 in the humanized Fab was also adjusted in shape and topography. The observed similarities in the functional binding activities of the two molecules can be rationalized by limited induced fit adjustments in their structures on antigen binding. While not perfect replicas, the two structures are testimonials to the progress in making high affinity monoclonal antibodies safe for human use.

Amino Acid Sequence↗