title: Origin and prediction of free-solution interaction studies performed label-free.
abstract: Interaction/reaction assays have led to significant scientific discoveries in the biochemical, medical, and chemical disciplines. Several fundamental driving forces form the basis of intermolecular and intramolecular interactions in chemical and biochemical systems (London dispersion, hydrogen bonding, hydrophobic, and electrostatic), and in the past three decades the sophistication and power of techniques to interrogate these processes has developed at an unprecedented rate. In particular, label-free methods have flourished, such as NMR, mass spectrometry (MS), surface plasmon resonance (SPR), biolayer interferometry (BLI), and backscattering interferometry (BSI), which can facilitate assays without altering the participating components. The shortcoming of most refractive index (RI)-based label-free methods such as BLI and SPR is the requirement to tether one of the interaction entities to a sensor surface. This is not the case for BSI. Here, our hypothesis is that the signal origin for free-solution, label-free determinations can be attributed to conformation and hydration-induced changes in the solution RI. We propose a model for the free-solution response function (FreeSRF) and show that, when quality bound and unbound structural data are available, FreeSRF correlates well with the experiment (R(2)> 0.99, Spearman rank correlation coefficients >0.9) and the model is predictive within ∼15% of the experimental binding signal. It is also demonstrated that a simple mass-weighted dη/dC response function is the incorrect equation to determine that the change in RI is produced by binding or folding event in free solution.
authors.affiliation: Department of Chemistry, Vanderbilt University, Nashville, TN 37235; The Vanderbilt Institute for Chemical Biology, Vanderbilt University, Nashville, TN 37235; darryl.bornhop@vanderbilt.edu.
authors.forename: Darryl J
authors.initials: DJ
authors.lastname: Bornhop
authors.affiliation: Department of Chemistry, Vanderbilt University, Nashville, TN 37235; The Vanderbilt Institute for Chemical Biology, Vanderbilt University, Nashville, TN 37235; Department of Biomedical Engineering, Vanderbilt University, Nashville, TN 37235;
authors.forename: Michael N
authors.initials: MN
authors.lastname: Kammer
authors.affiliation: Department of Chemistry, Vanderbilt University, Nashville, TN 37235; The Vanderbilt Institute for Chemical Biology, Vanderbilt University, Nashville, TN 37235;
authors.forename: Amanda
authors.initials: A
authors.lastname: Kussrow
authors.affiliation: Department of Chemistry, Lehigh University, Bethlehem, PA 18015;
authors.forename: Robert A
authors.initials: RA
authors.lastname: Flowers
authors.affiliation: Department of Chemistry, Vanderbilt University, Nashville, TN 37235; The Vanderbilt Institute for Chemical Biology, Vanderbilt University, Nashville, TN 37235; Vanderbilt University Center for Structural Biology, Nashville, TN 37235.
authors.forename: Jens
authors.initials: J
authors.lastname: Meiler
mesh_terms: D006860:Hydrogen Bonding; D007368:Interferometry / Q000379:methods*; D008956:Models, Chemical; D011485:Protein Binding; D011487:Protein Conformation; D017510:Protein Folding; D012031:Refractometry; D012680:Sensitivity and Specificity; D012996:Solutions; D012997:Solvents; D014867:Water
chemical_list: D012996:Solutions; D012997:Solvents; D014867:Water
keywords: assay methodology; backscattering interferometry; conformation change; hydration change; molecular interactions