4 research outputs found
Conformational Dynamics and Exchange Kinetics of <i>N</i>‑Formyl and <i>N</i>‑Acetyl Groups Substituting 3‑Amino-3,6-dideoxy-α‑d‑galactopyranose, a Sugar Found in Bacterial O‑Antigen Polysaccharides
Three
dimensional shape and conformation of carbohydrates are important
factors in molecular recognition events and the <i>N</i>-acetyl group of a monosaccharide residue can function as a conformational
gatekeeper whereby it influences the overall shape of the oligosaccharide.
NMR spectroscopy and quantum mechanics (QM) calculations are used
herein to investigate both the conformational preferences and the
dynamic behavior of <i>N</i>-acetyl and <i>N</i>-formyl substituents of 3-amino-3,6-dideoxy-α-d-galactopyranose,
a sugar and substitution pattern found in bacterial O-antigen polysaccharides.
QM calculations suggest that the amide oxygen can be involved in hydrogen
bonding with the axial OH4 group primarily but also with the equatorial
OH2 group. However, an NMR <i>J</i> coupling analysis indicates
that the θ<sub>1</sub> torsion angle, adjacent to the sugar
ring, prefers an <i>ap</i> conformation where conformations
<180° also are accessible, but does not allow for intramolecular
hydrogen bonding. In the formyl-substituted compound <sup>4</sup><i>J</i><sub>HH</sub> coupling constants to the <i>exo</i>-cyclic group were detected and analyzed. A van’t Hoff analysis
revealed that the <i>trans</i> conformation at the amide
bond is favored by Δ<i>G</i>° ≈ –
0.8 kcal·mol<sup>–1</sup> in the formyl-containing compound
and with Δ<i>G</i>° ≈ – 2.5 kcal·mol<sup>–1</sup> when the <i>N</i>-acetyl group is the substituent.
In both cases the enthalpic term dominates to the free energy, irrespective
of water or DMSO as solvent, with only a small contribution from the
entropic term. The <i>cis</i>–<i>trans</i> isomerization of the θ<sub>2</sub> torsion angle, centered
at the amide bond, was also investigated by employing <sup>1</sup>H NMR line shape analysis and <sup>13</sup>C NMR saturation transfer
experiments. The extracted transition rate constants were utilized
to calculate transition energy barriers that were found to be about
20 kcal·mol<sup>–1</sup> in both DMSO-<i>d</i><sub>6</sub> and D<sub>2</sub>O. Enthalpy had a higher contribution
to the energy barriers in DMSO-<i>d</i><sub>6</sub> compared
to in D<sub>2</sub>O, where entropy compensated for the loss of enthalpy
Molecular Dynamics Simulations of Membrane–Sugar Interactions
It is well documented that disaccharides
in general and trehalose
(TRH) in particular strongly affect physical properties and functionality
of lipid bilayers. We investigate interactions between lipid membranes
formed by 1,2-dimyristoyl-<i>sn</i>-glycero-3-phosphocholine
(DMPC) and TRH by means of molecular dynamics (MD) computer simulations.
Ten different TRH concentrations were studied in the range <i>w</i><sub>TRH</sub> = 0–0.20 (w/w). The potential of
mean force (PMF) for DMPC bilayer–TRH interactions was determined
using two different force fields, and was subsequently used in a simple
analytical model for description of sugar binding at the membrane
interface. The MD results were in good agreement with the predictions
of the model. The net affinities of TRH for the DMPC bilayer derived
from the model and MD simulations were compared with experimental
results. The area per lipid increases and the membrane becomes thinner
with increased TRH concentration, which is interpreted as an intercalation
effect of the TRH molecules into the polar part of the lipids, resulting
in conformational changes in the chains. These results are consistent
with recent experimental observations. The compressibility modulus
related to the fluctuations of the membrane increases dramatically
with increased TRH concentration, which indicates higher order and
rigidity of the bilayer. This is also reflected in a decrease (by
a factor of 15) of the lateral diffusion of the lipids. We interpret
these observations as a formation of a glassy state at the interface
of the membrane, which has been suggested in the literature as a hypothesis
for the membrane–sugar interactions
Stochastic Modeling of Flexible Biomolecules Applied to NMR Relaxation. 2. Interpretation of Complex Dynamics in Linear Oligosaccharides
A computational stochastic approach is applied to the
description
of flexible molecules. By combining (i) molecular dynamics simulations,
(ii) hydrodynamics approaches, and (iii) a multidimensional diffusive
description for internal and global dynamics, it is possible to build
an efficient integrated approach to the interpretation of relaxation
processes in flexible systems. In particular, the model is applied
to the interpretation of nuclear magnetic relaxation measurements
of linear oligosaccharides, namely a mannose-containing trisaccharide
and the pentasaccharide LNF-1. Experimental data are reproduced with
sufficient accuracy without free model parameters
Protein Flexibility and Conformational Entropy in Ligand Design Targeting the Carbohydrate Recognition Domain of Galectin-3
Rational drug design is predicated on knowledge of the three-dimensional structure of the protein−ligand complex and the thermodynamics of ligand binding. Despite the fundamental importance of both enthalpy and entropy in driving ligand binding, the role of conformational entropy is rarely addressed in drug design. In this work, we have probed the conformational entropy and its relative contribution to the free energy of ligand binding to the carbohydrate recognition domain of galectin-3. Using a combination of NMR spectroscopy, isothermal titration calorimetry, and X-ray crystallography, we characterized the binding of three ligands with dissociation constants ranging over 2 orders of magnitude. <sup>15</sup>N and <sup>2</sup>H spin relaxation measurements showed that the protein backbone and side chains respond to ligand binding by increased conformational fluctuations, on average, that differ among the three ligand-bound states. Variability in the response to ligand binding is prominent in the hydrophobic core, where a distal cluster of methyl groups becomes more rigid, whereas methyl groups closer to the binding site become more flexible. The results reveal an intricate interplay between structure and conformational fluctuations in the different complexes that fine-tunes the affinity. The estimated change in conformational entropy is comparable in magnitude to the binding enthalpy, demonstrating that it contributes favorably and significantly to ligand binding. We speculate that the relatively weak inherent protein−carbohydrate interactions and limited hydrophobic effect associated with oligosaccharide binding might have exerted evolutionary pressure on carbohydrate-binding proteins to increase the affinity by means of conformational entropy