1. The interaction of bioactive peptides with an immobilized phosphatidylcholine monolayer
H Mozsolits, T H Lee, H J Wirth, P Perlmutter, M I Aguilar Biophys J. 1999 Sep;77(3):1428-44. doi: 10.1016/S0006-3495(99)76991-2.
The interaction of three bioactive peptides, bombesin, beta-endorphin, and glucagon with a phosphatidylcholine monolayer that was immobilized to porous silica particles and packed into a stainless steel column cartridge, has been studied using dynamic elution techniques. This immobilized lipid monolayer provides a biophysical model system with which to study the binding of peptides to a lipid membrane. In particular, the influence of temperature and methanol concentration on the affinity of each peptide for the immobilized lipid surface was assessed. For all test peptides, nonlinear retention plots were observed at all temperatures that contrasted sharply with the simple linear plots observed for the small unstructured control molecules N-acetyltryptophanamide and diphenylalanine. An analysis of the thermodynamics of the interaction of peptides with the immobilized monolayer was also carried out. The results revealed that while the peptides interacted with the monolayer predominantly through hydrophobic interactions, the relative contribution of DeltaH(assoc)(O) and DeltaS(assoc)(O) to the overall free energy of association was dependent on the temperature and methanol concentration. In particular, it was evident that under most conditions, the binding of the peptides to the immobilized lipid monolayer was enthalpy-driven, i.e., mediated by nonclassical hydrophobic interactions. Significant band-broadening and asymmetric and split peaks were also observed for bombesin, beta-endorphin, and glucagon at different temperatures and methanol concentrations. These changes in affinity and peak shape are consistent with the formation of multiple conformational species during the interaction of these peptides with the lipid monolayer. In addition, the binding behavior of the three test peptides on an n-octylsilica surface that lacked the phospho headgroups of the phospholipid was significantly different from that observed with the immobilized phosphatidylcholine surface, indicating a specificity of interaction between the peptides and the lipid surface. Overall, these experimental results demonstrate that the biomimetic phosphatidylcholine monolayer provides a stable and sensitive system with which to explore the molecular mechanism of peptide conformational changes during membrane interactions.
2. High-performance liquid chromatography of amino acids, peptides and proteins. CXV. Thermodynamic behaviour of peptides in reversed-phase chromatography
A W Purcell, M I Aguilar, M T Hearn J Chromatogr. 1992 Feb 28;593(1-2):103-17. doi: 10.1016/0021-9673(92)80274-x.
The thermodynamic behaviour of three peptides, bombesin, beta-endorphin and glucagon, was studied under reversed-phase high-performance liquid chromatographic conditions. Experimental data related to the interactive surface contact area (S values) and solute affinity (log k0) were derived over a range of temperatures between 5 and 85 degrees C. These experimental conditions allowed changes in the secondary structure of the solute to be monitored. The influence of the nature of the stationary phase ligand on the relative conformational stability of the three peptides was analysed by acquiring data with n-octadecyl silica (C18) and n-butyl silica (C4) sorbents. Values for the relative changes in entropy and enthalpy associated with the interactive process were also determined. The results provide further insight into the factors involved with the stabilization of secondary structure and the mechanism of the interaction of peptides with hydrophobic surfaces.
3. Investigations into the thermodynamics of polypeptide interaction with nonpolar ligands
M T Hearn, G Zhao Anal Chem. 1999 Nov 1;71(21):4874-85. doi: 10.1021/ac990028x.
In this paper, we describe a general procedure to evaluate the thermodynamics of the interaction between polypeptides and hydrophobic ligands in the presence of aquo-organic solvent mixtures. These studies address experimental requirements for the determination of the linear free energy relationships, derivation of partition coefficients or other extrathermodynamic parameters such as contact areas, or assessment of the conformational changes that may occur when polypeptides or proteins interact with immobilized nonpolar ligands. Not unexpectedly from thermodynamic arguments, the trends and magnitudes of free energy parameters, such as the enthalpy of association, as previously derived in many studies from gradient elution reversed-phase high-performance liquid chromatographic (RP-HPLC) measurements are often different from the data for the same parameters derived from equilibrium binding or microcalorimetric determinations.
