Thermodynamic Stability and Flexibility Characteristics of Antibody Fragment Complexes

Abstract

Free energy landscapes, backbone flexibility and residue-residue couplings for being co-rigid or co-flexible are calculated from the minimal Distance Constraint Model (mDCM) on an exploratory dataset consisting of VL, scFv and Fab antibody fragments. Experimental heat capacity curves are reproduced markedly well, and an analysis of quantitative stability/flexibility relationships (QSFR) is applied to a representative VL domain and several complexes in the scFv and Fab forms. Global flexibility in the denatured ensemble typically decreases in the larger complexes due to domain-domain interfaces. Slight decreases in global flexibility also occur in the native state of the larger fragments, but with a concurrent large increase in correlated flexibility. Typically, a VL fragment has more co-rigid residue pairs when isolated compared to the scFv and Fab forms, where correlated flexibility appears upon complex formation. This context dependence on residue-residue couplings in the VL domain across length scales of a complex is consistent with the evolutionary hypothesis of antibody maturation. In comparing two scFv mutants with similar thermodynamic stability, local and long-ranged changes in backbone flexibility are observed. In the case of anti-p24 HIV-1 Fab, a variety of QSFR metrics were found to be atypical, which includes comparatively greater co-flexibility in the VH domain and less co-flexibility in the VL domain. Interestingly, this fragment is the only example of a polyspecific antibody in our dataset. Finally, the mDCM method is extended to cases where thermodynamic data is incomplete, enabling high throughput QSFR studies on large numbers of antibody fragments and their complexes.

JED: a Java Essential Dynamics Program for comparative analysis of protein trajectories

“…Watching movies at different time scales gives a sense of the effects of small and large amplitude motions (see Additional file 3 for movies of essential motions of the ScFv protein over different windows). In this case, the movies show the mutation rigidifies nearby residues in corroboration with our previous results [34]. To our knowledge, visualizing combination of modes within user-specified time scale windows offers a unique functionality/tool for researchers.…”

Dynamics and thermodynamic properties of CXCL7 chemokine

“…The mDCM parameters are u = −1.72 ± 0.26 kcal/mol, v = −0.47 ± 0.19 kcal/mol and δ gp = 1.20 ± 0.04 for the monomer, and u = −1.44 ± 0.27 kcal/mol, v = −0.11 ± 0.20 kcal/mol and δ gp = 1.43 ± 0.23 for the dimer. These parameters indicate that the native state of the dimer has greater conformational entropy per residue than that of the monomer (as seen before packing is not as strong,) while the destabilizing effect of H‐bonding from protein to solvent plays a lesser role. The maximum heat capacity, Cp max , of CXCL7 dimer is more than twice that of the monomers [Fig.…”

Novel Ricin Subunit Antigens With Enhanced Capacity to Elicit Toxin-Neutralizing Antibody Responses in Mice

“…Three empirical parameters that reflect solvent interactions are adjusted to fit to experimental DSC measurements. Since technical details can be found elsewhere 41–50 , only the procedure used to generate the results reported here are described.…”

“…Given an input protein structure at the all-atom level, the mDCM generates an ensemble of constraint topologies characterized by an intensive global flexibility order parameter defined as the number of independent degrees of freedom per residue. After the three empirical parameters are obtained following established protocols 47,50 , the free energy landscape is calculated as a function of temperature and global flexibility order parameter 48 . At temperature, T peak , where heat capacity is a maximum, a thermodynamic average for quantitative stability/flexibility relationships (QSFR) is taken in the native and transition state sub-ensembles, and these QSFR characteristics are compared between RiVax and its mutants.…”