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D C Wiley

Publications and source records attributed to D C Wiley.

At least 163 records · Page 9Linked to original sources
PubMed1988

Conformational aspects of the acid-induced fusion mechanism of influenza virus hemagglutinin. Circular dichroism and fluorescence studies.

Circular dichroism and tryptophan fluorescence spectroscopy have been used to investigate the structures of the influenza virus membrane glycoprotein hemagglutinin, acid-treated hemagglutinin, and fragments of hemagglutinin derived by proteolysis. The conformational change in hemagglutinin which occurs at the pH of membrane fusion (pH 5-6) was associated with a significant change of the environment of tyrosine residues, a change in the environment of tryptophan residues, but no changes in secondary structure. Tryptic digestion of the hemagglutinin in its low pH conformation which releases one of the subunit polypeptides (HA1) caused minimal changes in tyrosine and tryptophan environments but a small secondary structural change in HA1. The secondary structure of the remainder of the molecule (HA2) was very similar to that predicted from the known x-ray crystallographic structure of the native molecule. However, fluorescence spectroscopy indicated a tertiary change in structure in the coiled coil of alpha-helices which form the fibrous central stem of the molecule. These results are consistent with a conformational change required for membrane fusion which involves a decrease of HA1/HA1, HA1/HA2 interactions and changes in tertiary structure not accompanied by changes in secondary structure.

Circular Dichroism↗
PubMed1988

Studies on the structure of the influenza virus haemagglutinin at the pH of membrane fusion.

At the pH required to trigger the membrane fusion activity of the influenza virus haemagglutinin (HA) the soluble ectodomain of the molecule, BHA, which is released from virus by bromelain digestion, aggregates into rosettes. Analyses of soluble proteolytic fragments derived from the rosettes indicated that aggregation is mediated by association of the conserved hydrophobic amino-terminal region of BHA2, the smaller glycopolypeptide component of each BHA subunit. Further analyses of the structure of the soluble fragments and of HA in its low pH conformation by electron microscopy, spectroscopy and in crosslinking experiments showed that, although the membrane distal globular domains lose their trimer structure at the pH of fusion, the central fibrous stem of the molecule remains trimeric and assumes a more stable conformation. The increase in length of BHA2 at low pH observed microscopically appears to result from movement of the amino-terminal region to the membrane proximal end of the molecule and in virus incubated at low pH the amino terminus may insert into the virus membrane. The consequences of these possibilities for the mechanism of membrane fusion are discussed.

Bromelains↗
PubMed1988

Membrane fusion by peptide analogues of influenza virus haemagglutinin.

We have studied the interactions of synthetic peptides corresponding to the sequence of the amino terminus of the HA2 subunit of influenza virus haemagglutinin with artificial lipid membranes. The peptides could fuse cholesterol-free liposomes at neutral as well as acid pH; however, liposomes containing cholesterol could only be fused below pH 6. The fusion process caused leakage of aqueous liposomal contents. Peptides with amino acid substitutions had fusion properties similar to whole haemagglutinin molecules with the corresponding sequence changes. Non-fusogenic peptides still interacted with the membrane but did not cause leakage of liposomal contents. A correlation between the alpha-helical content of peptide and its fusogenicity was noted, but this was not absolute. The results reported here support suggestions for a role of the amino terminus of HA2 in virus-endosome fusion.

Hemagglutinins, Viral↗
PubMed1987

The receptor-binding and membrane-fusion properties of influenza virus variants selected using anti-haemagglutinin monoclonal antibodies.

A monoclonal antibody raised against X-31 influenza virus reacted with the majority of natural H3N2 viruses isolated between 1968 and 1982. A number of variants of X-31 and of a receptor-binding mutant of X-31 were selected by the antibody during virus replication in eggs and MDCK cells. Antibody-binding assays indicated that the viruses selected were not antigenic variants and analyses using derivatized erythrocytes showed that their receptor-binding properties differed from those of the parent viruses. The amino acid substitutions in the variants were all located in the vicinity of the receptor-binding site and the structural consequences are discussed in relation to the three-dimensional structure of X-31 HA. In addition all of the variants fused membranes at higher pH than wild-type virus indicating that structural modifications in the distal globular region of HA influence the low pH-induced conformational change required for membrane fusion.

Animals↗
PubMed1987

Receptor-binding characteristics of monoclonal antibody-selected antigenic variants of influenza virus.

Erythrocytes modified to different extents with periodate were used in hemagglutination assays to investigate the binding properties of antihemagglutinin monoclonal antibody-selected antigenic variants of X-31 influenza virus. The results allowed differentiation of groups of variants and are discussed in relation to the nature of the amino acid substitutions in the variant hemagglutinins and their molecular locations relative to the receptor-binding site.

Antibodies, Monoclonal↗
PubMed1987

Antigenic and amino acid sequence analysis of the variants of H1N1 influenza virus in 1986.

Since their reintroduction to human populations in 1977, influenza A viruses of the H1N1 subtype have undergone antigenic drift. Recently a distinct antigenic variant, A/Singapore/6/86, has been almost exclusively isolated internationally, and the antigenic properties and amino acid sequence of its haemagglutinin have been determined and compared with those of the haemagglutinins of other H1N1 viruses, in particular A/Chile/1/83. Fourteen amino acid sequence differences are detected between the HA1 components of these two viruses, ten of which are different from equivalent residues in the haemagglutinins of all H1N1 viruses isolated between 1982 and 1983, and seven of which are novel in the haemagglutinins of all H1N1 viruses sequenced to date. The results are discussed in relation to the three-dimensional structure of the haemagglutinin and the location of the previously defined antigenically important regions.

Amino Acid Sequence↗
PubMed1986

Protein, DNA, and virus crystallography with a focused imaging proportional counter.

A set of programs has been developed for rapid collection of x-ray intensity data from protein and virus crystals with a commercially available two-dimensional focused geometry electronic detector. The detector is compact and portable, with unusually high spatial resolution comparable to that used in oscillation photography. It has allowed x-ray data collection on weakly diffracting crystals with large unit cells, as well as more conventional "diffractometer-quality" crystals. The quality of the data is compared with that from oscillation photography and automated diffractometry in the range of unit cells from 96.3 to 383.2 angstroms. Isomorphous and anomalous difference Pattersons, based on detector data, are shown for a variable surface glycoprotein mercury derivative and for a repressor-DNA bromine derivative, which has been solved at 7 angstroms with detector data only.

Computers↗
PubMed1986

Studies of influenza haemagglutinin-mediated membrane fusion.

A resonance energy transfer assay of membrane fusion developed by P. S. Uster and D. W. Deamer (Biochemistry 24, 1-8 (1985)) was used in a study of influenza haemagglutinin-mediated fusion. The characteristics of fusion and haemolysis by X-31 (H3N2) virus, a number of mutants of X-31 which fuse membranes at higher pH, and purified haemagglutinins released from virus particles either by detergent dissociation or by bromelain digestion were compared with particular regard to pH and temperature dependence. The finding that membrane fusion activity, haemolysis, and changes in haemagglutinin conformation covary with pH and temperature provide support for the role of haemagglutinin in fusion and are discussed in relation to the stability of its structure.

Antibodies, Viral↗
PubMed1986

Conformational changes in the hemagglutinin of influenza virus which accompany heat-induced fusion of virus with liposomes.

Incubation of X-31 influenza virus at 62 degrees results in changes in hemagglutinin structure which cause the virus to fuse with liposomes at neutral pH. Mutants of X-31 which contain different hemagglutinins and as a consequence fuse with liposomes at higher pH than wild-type virus also fuse at a lower temperature. The heat-induced changes in hemagglutinin structure were monitored by CD, fluorescence spectroscopy, electron microscopy, and proteolytic digestion and compared with the characteristic changes which occur at low pH required for hemagglutinin-mediated membrane fusion at physiological temperature. The results indicate that the heat-induced changes in hemagglutinin which allow fusion activity result in partial denaturation of the molecule. By comparison the changes in structure required for fusion at low pH and normal temperature are specifically restricted.

Hemagglutinins, Viral↗
PubMed1986

Comparative analyses of the specificities of anti-influenza hemagglutinin antibodies in human sera.

Estimates of the variety of specificities of anti-influenza hemagglutinin antibodies in postinfection human sera taken between 1969 and 1971 and in 1978 were made by using Fab fragments of defined monoclonal antibodies in competitive virus-binding assays. The results obtained with the sera taken between 1969 and 1971 indicated that different sera contained antibodies with different ranges of specificities, whereas the 1978 sera mainly contained a broad range of antibodies. The results are discussed in relation to the mechanism of antigenic drift in influenza virus, the commonly observed antigenic heterogeneity of influenza virus isolates, and the efficacy of antiinfluenza vaccination.

Antibodies, Viral↗
PubMed1986

Biochemical and functional analyses of a secreted H-2Ld molecule.

A truncated H-2Ld gene was constructed by deleting the transmembrane and cytoplasmic exons. The truncated H-2Ld gene was introduced into mouse L cells using the thymidine kinase gene as a selectable marker. Transformants were isolated and screened for the presence of truncated H-2Ld antigen. The truncated H-2Ld gene product was present in both the cytoplasm and culture medium, but not on the cell surface. The truncated H-2Ld antigen was stable in culture medium for at least 9 h and was secreted into the medium at a rate similar to the kinetics with which complete H-2 antigens reach the cell surface. Transformants expressing the truncated H-2Ld molecule were not recognized by cytotoxic T lymphocytes specific for the H-2Ld antigen.

Animals↗
PubMed1985

Crystallization and X-ray diffraction studies on the histocompatibility antigens HLA-A2 and HLA-A28 from human cell membranes.

The human histocompatibility antigens HLA-A and HLA-B are polymorphic cell surface glycoproteins encoded by the major histocompatibility complex. These molecules are the major targets for the immune response during tissue transplantation. They are recognized by cytolytic T-lymphocytes during the immune response against virally infected cells, and have been linked to variations in susceptibility to human autoaggressive and neoplastic diseases. To permit a description of the sites of interaction with alloantisera and T-cell receptors, we have begun a three-dimensional structure determination of HLA-A. We report the isomorphous cyrstallization of two antigenic specificities of papain-solubilized HLA-A, A2 and A28. Isoelectric focusing indicates that the well-ordered crystals incorporate the sialic acid microheterogeneity of the oligosaccharides. Crystallographic evidence indicates that the HLA-A molecule has an approximate 2-fold rotational symmetry axis which, combined with biochemical data, suggests that the domains of the molecule are paired alpha 1 to alpha 2 and alpha 3 to beta 2-microglobulin. This domain organization is similar to the arrangement of domains in the Fab and Fc fragments of immunoglobulins.

Cell Membrane↗
PubMed1985

Host-mediated selection of influenza virus receptor variants. Sialic acid-alpha 2,6Gal-specific clones of A/duck/Ukraine/1/63 revert to sialic acid-alpha 2,3Gal-specific wild type in ovo.

Human and animal influenza A isolates of the H3 serotype preferentially bind SA alpha 2,6Gal or SA alpha 2,3Gal linkages (where SA represents sialic acid), respectively, on cell-surface sialyloligosaccharides. Previously, we have demonstrated selection of SA alpha 2,3Gal-specific receptor variants of several human viruses which differed from the parent viruses by a single amino acid at residue 226 of the hemagglutinin which is located in the receptor binding pocket (Rogers, G. N., Paulson, J.C., Daniels, R.S., Skehel, J.J., Wilson, I.A., and Wiley, D.C. (1983) Nature 304, 76-78). In this report, the selection in the reverse direction was accomplished starting with a SA alpha 2,3Gal-specific avian virus, A/duck/Ukraine/1/63 (H3N7), yielding SA alpha 2,6Gal-specific variants that exhibit the receptor binding properties characteristic of the human isolates. Selection was again mediated at residue 226 of the hemagglutinin, in this case changing from Gln in the parent virus to Leu in the variants. Although the SA alpha 2,6Gal-specific avian virus variants were stable to passage in MDCK cells, they exhibited dramatic reversion to the SA alpha 2,3Gal-specific phenotype of the parent virus during a single passage in chicken embryos. This was in contrast to the SA alpha 2,6Gal-specific human virus isolates which were stable to passage in both hosts. The reversion of the avian virus variants in eggs provides compelling evidence for host-mediated selection of influenza virus receptor variants.

Animals↗
PubMed1985

Fusion mutants of the influenza virus hemagglutinin glycoprotein.

The influenza virus hemagglutinin (HA) mediates viral entry into cells by a low pH induced membrane-fusion event in endosomal vesicles. Mutant viruses with altered pH dependence for both hemolysis and the HA conformational change required for fusion were selected for their ability to grow in cells treated with amantadine hydrochloride, which raises the endosomal pH. The amino acid sequence and three-dimensional location of 19 substitutions on the HA are reported. The mutations fall into two groups, one that results in the destabilization of the pH 7.0 location of the hydrophobic N-terminal HA2 peptide, and a second that results in the alteration of intersubunit contacts, suggesting a large distortion or disruption of these contacts in the "fusion-active" conformation.

Amantadine↗
PubMed1985

Amino acid sequences of haemagglutinins of influenza viruses of the H3 subtype isolated from horses.

The amino acid sequence of the haemagglutinin of A/equine/Miami/63 (H3N8), the prototype influenza virus of the H3 subtype from horses, is deduced from the nucleotide sequence of virus RNA and compared with the sequences of haemagglutinins of viruses of this subtype isolated from humans [X-31 (H3N2)] and from birds [A/duck/Ukraine/63 (H3N8)] and with the sequence of the haemagglutinin of A/equine/Fontainebleau/79 (H3N8) a virus isolated from a recent outbreak of equine influenza. The amino acid sequence differences detected are discussed with reference to the structure of the molecules, their antigenicity and antigenic drift in influenza viruses viruses isolated from horses.

Amino Acid Sequence↗
PubMed1985

Antigenic and amino acid sequence analyses of influenza viruses of the H1N1 subtype isolated between 1982 and 1984.

Since the reintroduction of influenza viruses of the H1N1 subtype into the human population in 1977, antigenically distinguishable viruses have been isolated each year from outbreaks of respiratory disease in young people. In this paper the antigenic properties of viruses representative of isolates made in 1982, 1983, and 1984 are compared, and the amino acid sequences of their haemagglutinins are presented. The results provide a complete description of the 1982 and 1983 H1N1 influenza virus haemagglutinins and allow correlation of specific amino acid sequence differences with antigenic differences.

Amino Acid Sequence↗