Small solutes affect protein and nucleic acid processes because of favorable or unfavorable chemical interactions of the solute with the biopolymer surface exposed or buried in the process. Large solutes also exclude volume and affect processes where biopolymer molecularity and/or shape changes. Here, we develop an analysis to separate and interpret or predict excluded volume and chemical effects of a flexible coil polymer on a process. We report a study of the concentration-dependent effects of the full series from monomeric to polymeric PEG on intramolecular hairpin and intermolecular duplex formation by 12-nucleotide DNA strands. We find that chemical effects of PEG on these processes increase in proportion to the product of the amount of DNA surface exposed on melting and the amount of PEG surface that is accessible to this DNA, and these effects are completely described by two interaction terms that quantify the interactions between this DNA surface and PEG end and interior groups. We find that excluded volume effects, once separated from these chemical effects, are quantitatively described by the analytical theory of Hermans, which predicts the excluded volume between a flexible polymer and a rigid molecule. From this analysis, we show that at constant concentration of PEG monomer, increasing PEG size increases the excluded volume effect but decreases the chemical interaction effect, because in a large PEG coil a smaller fraction of the monomers are accessible to the DNA. Volume exclusion by PEG has a much larger effect on intermolecular duplex formation than on intramolecular hairpin formation.m-value | macromolecular crowding | PEG | polymer excluded volume S olutes affect noncovalent biopolymer assembly processes such as folding, complex formation, and crystallization because the effect of the solute on the chemical potential of the products differs from its effect on the chemical potential of the reactants. Solute effects have been used to great advantage by biochemists for many years to optimize conditions for biochemical assays and to study the thermodynamics of protein folding and nucleic acid helix formation. A solute effect on a process is quantified by its "m-value," the derivative of the observed standard free energy change for the process with respect to the solute concentration, related to solute derivatives of reactant and product chemical potentials (μ 2 ) by Eq. 1:where component 3 is a perturbing solute and products (p) and reactants (r) are both considered to be component 2 (1). K obs is the equilibrium quotient of product concentrations to reactant concentrations, and K γ is the corresponding quotient of molar activity coefficients (γ 2 ). The last equality follows if m 2 is highly dilute and if m 3 is in significant excess over m 2 (2). For melting of a nucleic acid hairpin helix (h) to a denatured strand (s),and for melting of a duplex (d) to two strands (s 1 , s 2 ),If Δμ 2;3 is negative, then addition of solute either lowers the chemical potential of product moreso than reactant or r...

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