| 1. |
- Akerström, B, et al.
(författare)
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Binding properties of protein Arp, a bacterial IgA-receptor
- 1991
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Ingår i: Molecular Immunology. - 0161-5890. ; 28:4-5, s. 57-349
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Tidskriftsartikel (refereegranskat)abstract
- A cell surface receptor that binds to the Fc region of IgA is expressed by certain strains of group A streptococci. The physico-chemical properties and binding characteristics of this receptor, called protein Arp, were studied. Like bacterial receptors that bind IgG, protein Arp has an elongated shape and no disulfide bonds. The affinity constant of protein Arp for three different molecular forms of IgA was determined, and was found to be more than ten-fold higher for serum IgA than for two complexed forms of IgA: secretory IgA and IgA bound to alpha 1-microglobulin. Cleavage of protein Arp with CNBr resulted in a peptide corresponding to the region located outside the cell wall, except for the N-terminal 52 amino acids. This CNBr-fragment did not bind IgA, which strongly suggests that the IgA-binding region of protein Arp is located in the N-terminal part of the molecule. In addition to the binding of IgA, protein Arp also binds to IgG weakly. The pH-dependence of these two types of binding is different, with maximal binding of IgA at neutral pH (5-7) and maximal binding of IgG at acidic pH (3-5). Both for IgA and IgG, protein Arp shows strong specificity for immunoglobulins of human origin.
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| 2. |
- Dueñas, Marta, et al.
(författare)
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Selection of phage displayed antibodies based on kinetic constants
- 1996
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Ingår i: Molecular Immunology. - : Elsevier BV. - 0161-5890. ; 33:3, s. 279-285
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Tidskriftsartikel (refereegranskat)abstract
- The display of antibody fragments on the surface of filamentous bacteriophages and the selection of binders from antibody libraries have provided powerful tools to generate human antibodies. We reported recently a new concept (SAP system) for the selection of specific phages by linking antigenic recognition and phage replication, using a soluble fusion protein containing the antigen and a fragment of the M13 coat protein 3. In this investigation, a model library has been composed using six different antibody fragments which were characterized individually regarding their k(ass), k(diss) and K(a). All Feb fragments were specific for a 15 amino acid region of the V3 loop of gp120 (HIV-1). We demonstrated that the SAP system could discriminate between the kinetic parameters of each clone, using different selection strategies. Phages expressing high affinity clones were selected preferentially using low doses of antigen but clones of lower affinity also could be selected by increasing the antigen concentration or using a preselection procedure. Phages expressing antibody fragment with high association or low dissociation rate constants were retrieved by utilizing short contact times between antigen and antibody or antigen-chase conditions.
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| 3. |
- Ohlin, Mats, et al.
(författare)
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Characteristics of human antibody repertoires following active immune responses in vivo
- 1996
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Ingår i: Molecular Immunology. - : Elsevier BV. - 0161-5890. ; 33:7-8, s. 583-592
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Tidskriftsartikel (refereegranskat)abstract
- Possibilities to develop human monoclonal antibody specificities have recently been much facilitated by improvements of human hybridoma technology but even more so by the emerging phage-display technique. However, until recently very little has been known about the characteristics at the molecular level of the induced, T cell-dependent human antibody response, frequently targeted by these techniques. Rather, the major part of available sequence information has been related to tumor-derived or autoreactive antibodies. We have now investigated high affinity, monospecific, human antibody repertoires as developed by hybridoma technology. The VH region gene usage among such in vivo-induced repertoires is in only some respects similar to that found in the total B cell population. A limited number of heavy-chain variable segment loci account for the majority of all induced antibodies. A particular VH gene locus (4-34) frequently employed by peripheral B cells and associated with autoreactive antibodies was rarely used by the induced repertoire. Furthermore, in particular antigen systems, V region usage differs from the total available repertoire, and heavy-chain CDR3 is generally longer among antibodies induced against foreign protein antigens than in the average B cell population. Light-chain gene usage is often restricted to just a few dominant genes frequently found among B cells in general. In comparison, variable regions derived by phage-display technology in some antigen systems display even longer heavy-chain CDR3 than hybridoma-derived antibodies. This technique also appears to select a different set of germline genes preferentially (both with respect to VH and JH) as compared to hybridoma technology. In summary, the T cell-dependent antibody response against foreign antigens appears to differ from the average circulating B cell in several ways, and thus does not seem to represent a random selection of the available repertoire.
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| 4. |
- Ohlin, Mats, et al.
(författare)
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Insertions and deletions in hypervariable loops of antibody heavy chains contribute to molecular diversity
- 1998
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Ingår i: Molecular Immunology. - 0161-5890. ; 35:4, s. 233-238
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Tidskriftsartikel (refereegranskat)abstract
- Antibody diversity, a molecular feature which allows these proteins to specifically interact with a diverse set of targets, is created at the genetic level by a variety of means. These include germline gene segment recombination, junctional diversity and single basepair (bp) substitution. We here demonstrate that a human high affinity antibody specific for an exogenous protein antigen carry three amino acid residues immediately adjacent to the first hypervariable loop of the heavy chain. These additional residues are shown not to be encoded by the germline repertoire. We also describe the characteristics of insertions and deletions, not found in any known germline sequence, within the first and second hypervariable loops of other previously described antibody-encoding genes. These findings demonstrate that insertions or deletions of entire codons provide yet another approach by which the human antibody repertoire is diversified in vivo. Since these major genetic modifications occur within or immediately adjacent to loops contributing to the antigen-binding site, they are likely to affect the binding properties of the mutated antibodies.
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| 5. |
- Ohlin, Mats, et al.
(författare)
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Light chain shuffling of a high affinity antibody results in a drift in epitope recognition
- 1996
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Ingår i: Molecular Immunology. - : Elsevier BV. - 1872-9142 .- 0161-5890. ; 33:1, s. 47-56
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Tidskriftsartikel (refereegranskat)abstract
- Human polyclonal and monoclonal antibodies against pathogens and toxins are potentially useful in the treatment of various diseases. A number of human monoclonal antibodies with protective capacity in vitro have been established by conventional hybridoma technology. However, with the development of phage-display technology, the possibility of specifically tailoring antigen-binding properties has improved substantially. We show here that the reactivity of a high affinity, virus-neutralizing human antibody against the AD-2 epitope of cytomegalovirus gB can be modified by introducing other Vkappa sequences together with the original VH sequence. The fine specificity, as determined by the requirement of particular amino acid residues in the epitope, is shifted in these new antibody fragments. It was also evident that the VH/Vkappa pairing was not promiscuous, since antibody fragments selected by phage display retained light chain sequences very similar to the original hybridoma-derived light chain, proving that a high affinity interaction was very dependent on a co-operativity between both variable domains. These findings show that phage display technology might modify the binding properties of pre-existing, high affinity antibodies.
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| 6. |
- Ohlin, Mats, et al.
(författare)
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Restricted variable region gene usage and possible rheumatoid factor relationship among human monoclonal antibodies specific for the AD-1 epitope on cytomegalovirus glycoprotein B
- 1994
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Ingår i: Molecular Immunology. - : Elsevier BV. - 1872-9142 .- 0161-5890. ; 31:13, s. 983-991
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Tidskriftsartikel (refereegranskat)abstract
- The nucleotide sequences of the variable region genes encoding five different human, high affinity antibodies, specific for the major neutralization determinant (AD-1) expressed by human cytomegalovirus glycoprotein B (gp58/116), have been determined. Three of the five heavy chain variable regions belonged to the small VHV-family, although they combined with a diverse set of light chains (V kappa IIIb, V lambda II and V lambda III). The other two antibodies belonged to VH-families III and IV. One of the VHV-family genes most likely originated from a previously unreported germline gene or allele, since it carries a nine nucleotide insert in framework 1. In addition, V lambda-genes showed variable homology (77-95%) to known germline sequences, while V kappa-genes showed high homology (approximately 98%) with their proposed germline origin. Despite the close homology of the V kappa IIIb-gene used to express one of the antibodies with its corresponding germline gene, the protein did not strongly express some idiotypes associated with this light chain family. There is, thus, no direct relation between the expression of these crossreactive idiotypes and the use of even modestly mutated light chains belonging to this V kappa-family, which has been implicated in the development of anti-idiotypic networks possibly inducing autoantibodies, such as rheumatoid factors.
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| 7. |
- Sabharwal, H, et al.
(författare)
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Blood group specific oligosaccharides from faeces of a blood group A breast-fed infant
- 1984
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Ingår i: Molecular Immunology. - : Elsevier BV. - 1872-9142 .- 0161-5890. ; 21:11, s. 1105-1112
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Tidskriftsartikel (refereegranskat)abstract
- Four different oligosaccharides were isolated from faeces collected from a blood group A, secretor, breast-fed infant. Three of these, GalNAc alpha 1-3[Fuc alpha 1-2]Gal beta 1-4Glc (A-tetrasaccharide), GalNAc alpha 1-3[Fuc alpha 1-2]Gal beta 1-4[Fuc alpha 1-3]Glc (A-pentasaccharide) and 1-3[Fuc alpha 1-4]GlcNAc beta 1-3Gal beta 1-4Glc (A-heptasaccharide) have previously found in urine, whereas GalNAc alpha 1-3[Fuc alpha 1-2]Gal beta 1-3GlcNAc beta 1-3Gal beta 1-4Glc (A-hexasaccharide) is a new compound. Structures were deduced by mass spectrometry of permethylated and N-trifluoroacetylated oligosaccharide alditols. The latter gave more structural information than the corresponding N-acetyl derivatives. The four oligosaccharides were tested for blood group A activity and all were found to inhibit the binding of anti-A antibody to blood group A substance.
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| 8. |
- Wingren, Christer, et al.
(författare)
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Antigen-binding sites dominate the surface properties of IgG antibodies
- 1995
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Ingår i: Molecular Immunology. - 1872-9142 .- 0161-5890. ; 32:11, s. 819-827
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Tidskriftsartikel (refereegranskat)abstract
- A new technique, liquid-liquid partition chromatography in an aqueous polyethylene glycol-dextran two-phase system, was used to detect differences in surface properties of antibodies with different antigen-binding sites. Employing well-characterized monoclonal IgG antibodies and Fab and Fc fragments thereof as well as chimeric IgG antibodies we found a remarkable relationship between structure of the antibody combining site and chromatographic behaviour. The surface properties of the IgG antibodies were dominated by those of its antigen-binding regions. In addition, our results indicated that the constant parts of the IgGs form similar scaffoldings, on to which CDRs of variable shapes and sizes are interspaced and constitute the major dominant differences in exposed surface properties.
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| 9. |
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| 10. |
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