A heterologous immunoglobulin chain recombinant carries a distinct site for dinitrophenyl and obeys the common hapten binding mechanism

Zidovetzki, Raphael; Blatt, Yoav; Pecht, Israel

doi:10.1021/bi00520a030

Cited by 12 publications

(8 citation statements)

References 41 publications

(41 reference statements)

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“…This conclusion is consistent with earlier observations (e.g. Zidovetzki et al, 1981;Davies and Metzger, 1983;Figini et al, 1994). Moreover, our results show that this flexibility is related to the degree of compactness of the interface and can affect binding affinity.…”

Section: Experimental Applicationssupporting

confidence: 94%

Unraveling the effect of changes in conformation and compactness at the antibody VL-VH interface upon antigen binding

et al. 1999

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We have analyzed conformational changes that occur at the interface between the light (V(L)) and heavy (V(H)) chains in antibody variable fragments upon binding to antigens. We wrote and applied the Tiny Probe program that computes the buried atomic contact surface area of three-dimensional structures to evaluate changes in compactness of the V(L)-V(H) interface between bound and unbound antibodies. We found three categories of these changes, which correlated with the size of the antigen. Upon binding, medium-sized nonprotein antigens cause an opening of the V(L)-V(H) interface (less compact), small antigens or haptens cause a closure of the interface (more compact), whereas large protein antigens have little effect on the compactness of the V(L)-V(H) interface. The largest changes in the atomic buried contact surface area at the V(L)-V(H) interface occur in residue pairs providing two 'shock absorbers' between the edge beta-strands of the V(L) and V(H) beta-sheets forming the antibody binding site. Importantly, the correlation between the size of antigens and conformational changes indicates that the V(L)-V(H) interface in antibodies plays a significant role in the antigen binding process. Furthermore, as the energy involved in such a motion is significant (up to 3 kcal/mol), these results provide a general mechanism for how residues distant from the combining site can significantly alter the affinity of an antibody for its antigen. Thus, mutations introduced at the V(L)-V(H) interface can be used to change antibody binding affinity with antigens. Due to the tightly packed V(L)-V(H) interface, the introduction of random mutations is not advisable. Rather our analysis suggests that concerted mutations of residues preceding CDRL2 and following CDRH3 or residues preceding CDRH2 and at the end of CDRL3 are most likely to alter or improve antigen binding affinity.

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Section: Experimental Applicationssupporting

confidence: 94%

Unraveling the effect of changes in conformation and compactness at the antibody VL-VH interface upon antigen binding

et al. 1999

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“…In contrast, fast kinetic measurements of antibody-antigen interactions (antibodies are the soluble form of the BCR) had been extensively performed in seminal studies by Israel Pecht's group (10,11). These studies provided consistent kinetic evidence that conformational changes are taking place in the variable loops of the antibodies induced by antigen binding.…”

mentioning

confidence: 99%

The TCR binding site does move

Schamel¹,

Reth²

2007

Proc. Natl. Acad. Sci. U.S.A.

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“…Since slightly different relative arrangements of VH and VL have been observed in the crystal structures of several antibodies and light-chain dimers (7,13,14), it is likely that they reflect a relative motion of the domains in solution. Results of kinetic and thermodynamic studies of hapten binding by mAbs have led to the proposal that VH-VL interdomain interactions are stabilized by binding (15,16), as seems to be the case in the Fv D1.3-HEL complex (7). These conformational changes are consistent with an "induced fit" mechanism, as described for other antibodies (2,17,18).…”

mentioning

confidence: 99%

Bound water molecules and conformational stabilization help mediate an antigen-antibody association.

Bhat

Bentley

Boulot

et al. 1994

Proc. Natl. Acad. Sci. U.S.A.

473

336

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We report the three-dimensional structures, at 1.8-A resolution, ofthe Fv fragment ofthe anti-hen egg white lysozyme antibody D1.3 in its free and antigen-bound forms.These structures reveal a role for solvent molecules in stabilzing the complex and provide a molecular basis for understanding the thermodynamic forces which drive the association reaction. Four water molecules are buried and others form a hydrogen-bonded network around the interface, bridging antigen and antibody. Comparison of the structures of free and bound Fv fragment of D1.3 reveals that several of the ordered water molecules in the free antibody combining site are retained and that additional water molecules link antigen and antibody upon complex formation. This salvation of the complex should weaken the hydrophobic effect, and the resulting large number of solvent-mediated hydrogen bonds, in conijunction with direct protein-protein interactions, should generate a significant enthalpic component. Furthermore, a stabilization of the relative mobilities of the antibody heavy-and light-chain variable domains and of that of the third complementaritydetermining loop of the heavy chain seen in the complex should generate a negative entropic contribution opposing the enthalpic and the hydrophobic (solvent entropy) effects. This structural analysis is consistent with measurements of enthalpy and entropy changes by titration calorimetry, which show that enthalpy drives the antigen-antibody reaction. Thus, the main forces stabilizing the complex arise from antigen-antibody hydrogen bonding, van der Waals interactions, enthalpy of hydration, and conformational stabilization rather than solvent entropy (hydrophobic) effects.X-ray crystallographic studies of several complexes of antigens with specific antibodies have revealed a high degree of complementarity between their interacting surfaces (reviewed in refs. 1 and 2). Water molecules have been identified at the interfaces of the Fab fragment of antibody D1.3 (Fab D1.3)-hen egg-white lysozyme (HEL) (3) and NC41-neuraminidase (4) complexes on the basis of structure determinations at 2.5-A resolution. Unfortunately, at such resolution, which is about the best which has been so far attained with conventional Fab fragments, the certainty with which ordered water molecules can be located is seriously limited (5). We have now determined the three-dimensional structure, at 1.8-A resolution, of the Fv fragment of monoclonal antibody (mAb) D1.3 (6, 7), Fv D1.3, consisting of only the variable domains ofthe heavy (VH) and light (VL) polypeptide chains and that of its complex with HEL, permitting a more detailed description of an antibody combining site in its free and antigen-bound states. These studies reveal both buried and exposed water molecules linking antigen and antibody and contributing to chemical complementarity between their interacting surfaces.An understanding of how antibodies react with antigens must involve the thermodynamics of the binding interaction. We have therefore experimentally de...

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A heterologous immunoglobulin chain recombinant carries a distinct site for dinitrophenyl and obeys the common hapten binding mechanism

Cited by 12 publications

References 41 publications

Unraveling the effect of changes in conformation and compactness at the antibody VL-VH interface upon antigen binding

Unraveling the effect of changes in conformation and compactness at the antibody VL-VH interface upon antigen binding

The TCR binding site does move

Bound water molecules and conformational stabilization help mediate an antigen-antibody association.

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