Biomedical subjects

F R Carbone

Publications and source records attributed to F R Carbone.

77 records · Page 5Linked to original sources

PubMed1987

The use of hydrophobic, alpha-helix-defined peptides in delineating the T cell determinant for pigeon cytochrome c.

The B10.A T cell proliferative response to pigeon cytochrome c is largely directed to a single site in the molecule located at the carboxyl terminus within the amino acid sequence of residues 81 to 104. This study uses the pigeon cytochrome c-specific T cell clone A.E7 and synthetic peptide analogs to clarify the role of certain residues within this sequence in T cell recognition. By using the helically constrained amino acid, alpha-aminoisobutyric acid, alternated with alanine in an amino-terminal leader sequence, we generated a series of molecules of similar length and alpha-helical conformation but which contain increasing lengths of the native sequence. By comparing the stimulatory ability of this series of peptides, we have clearly identified that the isoleucyl residue at position 95 in pigeon cytochrome c is essential for T cell recognition. This series, when compared with a series containing the same native sequences but without the leader sequence, also showed that the presence of the leader sequence has a general effect on enhancement of T cell recognition. An analysis of the conformational preferences of the peptides using circular dichroism indicated that all of the peptides with leader sequences have a strong preference for the alpha-helical conformation in nonpolar solvents. However, the introduction of helix-breaking residues into these peptides, with a concomitant measured reduction in alpha-helix, did not affect their recognition by clone A.E7. This implies that factors other than conformational stabilization are responsible for the full potency of these peptides. Binding studies to phospholipid vesicles indicated that residues in the leader sequence and in the amino terminus of segment 81-104 beyond residue 95 were important in increasing the ability of the antigens to bind to membranes. These results suggest that the capacity to bind to membranes may be a significant factor in the dose response of T cells to exogenously presented peptides.

Animals↗

PubMed1987

A new T helper cell specificity within the pigeon cytochrome c determinant 95-104.

A T cell hybridoma with unusual antigen specificity has been isolated from B10.BR mice immunized with pigeon cytochrome c. Using synthetic peptide analogues of the pigeon cytochrome c T cell determinant 81-104 we have determined that this hybridoma is specific for residues within the sequence 95-104. However, the mode by which this T cell hybridoma recognizes this determinant in the context of the I-Ek molecule differs from all previously described T cell clones in that it is not stimulated by the moth cytochrome c analogue 81-103.

Amino Acid Sequence↗

PubMed1987

Evidence for the role of a human intestinal adenovirus in the pathogenesis of coeliac disease.

We previously noted a region of amino acid sequence homology between A-gliadin, a major alpha-gliadin component known to activate coeliac disease, and the early region E1b protein of human adenovirus serotype 12 (Ad12), an adenovirus isolated from the human intestinal tract. In the present study sera from coeliac disease patients from the United Kingdom and the United States were assayed for neutralising antibody to Ad12 as evidence of past exposure to that virus and for antibody to synthetic peptides of A-gliadin from the region of shared sequence with the Ad12 E1b protein. Eighty nine per cent of untreated coeliac disease patients had evidence of previous Ad12 infection. There was also a significant increase in the prevalence of neutralising antibody to Ad12 among treated adults (33.3%) and children (30.8%) with coeliac disease compared with controls (0-12.8%) in the western USA and in London. There was no evidence for an increased prevalence of infection with a closely related adenovirus, adenovirus 18, or another enteric virus, Echovirus 11, among coeliac disease subjects. Additional studies documented that a region of A-gliadin that shares amino acid sequence homology with the adenovirus 12 E1b protein could be recognised as an antigenic determinant in active coeliac disease patients. Taken together, these data are compatible with the hypothesis that a viral protein may play a role in the pathogenesis of coeliac disease, perhaps by virtue of immunological cross reactivity between antigenic determinants shared by the viral protein and alpha-gliadins.

Adenoviridae Infections↗

PubMed1985

Two large immunogenic and antigenic myoglobin peptides and the effects of cyclisation.

Peptides corresponding to sequences (72-88) and (26-54) of beef myoglobin have been synthesised in their open-chain and cyclised forms (using a disulphide bridge) and tested for their antigenicity and immunogenicity. Antibodies raised to beef myoglobin bound to both peptides but more strongly to the 29-residue than to the 17-residue peptide. Cyclisation increased the antigenicity of the larger peptide. In this form the peptide competed much more strongly than in the uncyclised form for specific antibodies to beef myoglobin. The peptides are immunogenic in mice without being coupled to a protein carrier and produce antibodies which bind to beef myoglobin. Peptide (26-54) is the more immunogenic in producing a larger antibody titre to the parent myoglobin and cyclisation again enhances this property. The findings lend weight to the view that longer peptide sequences might be expected to favour the folded state, therefore binding more strongly to antibodies raised to the native protein and eliciting a population of antibodies which contain a larger proportion specific for that conformation. Cyclisation enhances antigenicity and immunogenicity presumably by decreasing the number of degrees of conformational freedom of a peptide without excluding native-like conformations.

Amino Acid Sequence↗

PubMed1985

Monoclonal antibodies to horse cytochrome c expressing four distinct idiotypes distribute among two sites on the native protein.

Antibody-secreting hybridoma cell lines produced from BALB/c mice that had been immunized with horse cytochrome c (cyt c) conjugated to hemocyanin yielded six hybridoma subclones that produced four monoclonal antibodies (mAb) with different patterns of cross-reactivity with a panel of evolutionarily variant cyt c. The recognition sites for three of these mAb lay in the same region of the intact molecule, because two of the mAb were sensitive to the amino acid residue present at sequence position 44, with one requiring threonine at position 47. The fourth mAb bound in another region of the molecule at a site that involves either residue 60 or residue 89. Synthetic peptides that included these residues did not react with these mAb, indicating that these sites may require interactions from noncontiguous regions of the molecule to bind antibody. The association constants for the interaction of the mAb with horse cyt c were very similar and of the order of 10(10) M-1. Specificity studies with anti-idiotypic sera and competition assays between mAb for binding to horse cyt c confirmed that the six positive hybridoma subclones produced from this fusion produced mAb that had one of these four distinct specificities. The idiotypes of these four mAb were serologically distinct, and were derived from Vh genes of the J558 family.

Animals↗