High Affinity Antigen Recognition of the Dual Specific Variants of Herceptin Is Entropy-Driven in Spite of Structural Plasticity

The antigen-binding site of Herceptin, an anti-human Epidermal Growth Factor Receptor 2 (HER2) antibody, was engineered to add a second specificity toward Vascular Endothelial Growth Factor (VEGF) to create a high affinity two-in-one antibody bH1. Crystal structures of bH1 in complex with either antigen showed that, in comparison to Herceptin, this antibody exhibited greater conformational variability, also called “structural plasticity”. Here, we analyzed the biophysical and thermodynamic properties of the dual specific variants of Herceptin to understand how a single antibody binds two unrelated protein antigens. We showed that while bH1 and the affinity-improved bH1-44, in particular, maintained many properties of Herceptin including binding affinity, kinetics and the use of residues for antigen recognition, they differed in the binding thermodynamics. The interactions of bH1 and its variants with both antigens were characterized by large favorable entropy changes whereas the Herceptin/HER2 interaction involved a large favorable enthalpy change. By dissecting the total entropy change and the energy barrier for dual interaction, we determined that the significant structural plasticity of the bH1 antibodies demanded by the dual specificity did not translate into the expected increase of entropic penalty relative to Herceptin. Clearly, dual antigen recognition of the Herceptin variants involves divergent antibody conformations of nearly equivalent energetic states. Hence, increasing the structural plasticity of an antigen-binding site without increasing the entropic cost may play a role for antibodies to evolve multi-specificity. Our report represents the first comprehensive biophysical analysis of a high affinity dual specific antibody binding two unrelated protein antigens, furthering our understanding of the thermodynamics that drive the vast antigen recognition capacity of the antibody repertoire

Entropy-Enthalpy Compensation May Be a Useful Interpretation Tool for Complex Systems Like Protein-DNA Complexes: An Appeal to Experimentalists

In various chemical systems enthalpy-entropy compensation (EEC) is a well-known rule of behavior, although the physical roots of it are still not completely understood. It has been frequently questioned whether EEC is a truly physical phenomenon or a coincidence due to trivial mathematical connections between statistical-mechanical parameters - or even simpler: A phantom effect resulting from the misinterpretation of experimental data. Here, we review EEC from a new standpoint using the notion of correlation which is essential for the method of factor analysis, but is not conventional in physics and chemistry. We conclude that the EEC may be rationalized in terms of hidden (not directly measurable with the help of the current experimental set-up) but physically real factors, implying a Carnot-cycle model in which a micro-phase transition (MPT) plays a crucial role. Examples of such MPTs underlying physically valid EEC should be typically cooperative processes in supramolecular aggregates, like changes of structured water at hydrophobic surfaces, conformational transitions upon ligand-biopolymer binding, and so on, so forth. The MPT notion could help rationalize the occurrence of EEC in connection with hydration and folding of proteins,enzymatic reactions, functioning of molecular motors, DNA de- and rehybridization, as well as similar phenomena.Comment: 8 pages, 2 Figures, Submitted for publicatio

Thermodynamic pathways to genome spatial organization in the cell nucleus

The architecture of the eukaryotic genome is characterized by a high degree of spatial organization. Chromosomes occupy preferred territories correlated to their state of activity and, yet, displace their genes to interact with remote sites in complex patterns requiring the orchestration of a huge number of DNA loci and molecular regulators. Far from random, this organization serves crucial functional purposes, but its governing principles remain elusive. By computer simulations of a Statistical Mechanics model, we show how architectural patterns spontaneously arise from the physical interaction between soluble binding molecules and chromosomes via collective thermodynamics mechanisms. Chromosomes colocalize, loops and territories form and find their relative positions as stable hermodynamic states. These are selected by “thermodynamic switches” which are regulated by concentrations/affinity of soluble mediators and by number/location of their attachment sites along chromosomes. Our “thermodynamic switch model” of nuclear architecture, thus, explains on quantitative grounds how well known cell strategies of upregulation of DNA binding proteins or modification of chromatin structure can dynamically shape the organization of the nucleus

Physical Constraints and Functional Characteristics of Transcription Factor-DNA Interaction

We study theoretical ``design principles'' for transcription factor-DNA interaction in bacteria, focusing particularly on the statistical interaction of the transcription factors (TF's) with the genomic background (i.e., the genome without the target sites). We introduce and motivate the concept of `programmability', i.e. the ability to set the threshold concentration for TF binding over a wide range merely by mutating the binding sequence of a target site. This functional demand, together with physical constraints arising from the thermodynamics and kinetics of TF-DNA interaction, leads us to a narrow range of ``optimal'' interaction parameters. We find that this parameter set agrees well with experimental data for the interaction parameters of a few exemplary prokaryotic TF's. This indicates that TF-DNA interaction is indeed programmable. We suggest further experiments to test whether this is a general feature for a large class of TF's.Comment: 9 pages, 4 figures; revised version as published in PNA

The thermodynamics of general anesthesia

It is known that the action of general anesthetics is proportional to their partition coefficient in lipid membranes (Meyer-Overton rule). This solubility is, however, directly related to the depression of the temperature of the melting transition found close to body temperature in biomembranes. We propose a thermodynamic extension of the Meyer-Overton rule which is based on free energy changes in the system and thus automatically incorporates the effects of melting point depression. This model provides a quantitative explanation of the pressure reversal of anesthesia. Further, it explains why inflammation and the addition of divalent cations reduce the effectiveness of anesthesia.Comment: 7 pages, 2 figure