Energetics of intermolecular hydrogen bonds in a hydrophobic protein cavity

Abstract

This work explores the energetics of intermolecular H-bonds inside a hydrophobic protein cavity. Kinetic measurements were performed on the gaseous deprotonated ions (at the -7 charge state) of complexes of bovine β-lactoglobulin (Lg) and three monohydroxylated analogs of palmitic acid (PA): 3-hydroxypalmitic acid (3-OHPA), 7-hydroxypalmitic acid (7-OHPA), and 16-hydroxypalmitic acid (16-OHPA). From the increase in the activation energy for the dissociation of the (Lg + X-OHPA) 7- ions, compared with that of the (Lg + PA) 7- ion, it is concluded that the -OH groups of the X-OHPA ligands participate in strong (5 - 11 kcal mol -1) intermolecular H-bonds in the hydrophobic cavity of Lg. The results of molecular dynamics (MD) simulations suggest that the -OH groups of 3-OHPA and 16-OHPA act as H-bond donors and interact with backbone carbonyl oxygens, whereas the -OH group of 7-OHPA acts as both H-bond donor and acceptor with nearby side chains. The capacity for intermolecular H-bonds within the Lg cavity, as suggested by the gas-phase measurements, does not necessarily lead to enhanced binding in aqueous solution. The association constant (K a) measured for 7-OHPA [(2.3 ± 0.2) × 10 5 M -1 ] is similar to the value for the PA [(3.8 ± 0.1) × 10 5 M -1 ]; K a for 3-OHPA [(1.1 ± 0.3) × 10 6 M -1 ] is approximately three-times larger, whereas K a for 16-OHPA [(2.3 ± 0.2) × 10 4 M -1 ] is an order of magnitude smaller. Taken together, the results of this study suggest that the energetic penalty to desolvating the ligand -OH groups, which is necessary for complex formation, is similar in magnitude to the energetic contribution of the intermolecular H-bonds. [Figure not available: see fulltext.] © 2014 American Society for Mass Spectrometry.