ABSTRACT
Fluorophores linked to the glucose/galactose-binding protein (GGBP) are a promising class of glucose sensors with potential application in medical devices for diabetes patients. Several different fluorophores at different positions in the protein were tested experimentally so far, but a deeper molecular understanding of their function is still missing. In this work, we use molecular dynamics simulations to investigate the mechanism of glucose binding in the GGBP-Badan triple mutant and make a comparison to the GGBP wild-type protein. The aim is to achieve a detailed molecular understanding of changes in the glucose binding site due to the mutations and their effect on glucose binding. Free simulations give an insight into the changes of the hydrogen-bonding network in the active site and into the mechanisms of glucose binding. Additionally, metadynamics simulations for wild type and mutant unravel the energetics of binding/unbinding in these proteins. Computed free energies for the opening of the binding pocket for the wild-type and the mutant agree well with the experimental data. Further, the simulations also give an insight into the changes of the chromophore conformations upon glucose binding, which can help to understand fluorescence changes. Therefore, the molecular details unravelled in this work may support effective optimisation strategies for the construction of more efficient glucose sensors.
Subject(s)
Escherichia coli Proteins/metabolism , Glucose/metabolism , Monosaccharide Transport Proteins/metabolism , Binding Sites , Escherichia coli Proteins/chemistry , Escherichia coli Proteins/genetics , Fluorescent Dyes/chemistry , Glucose/chemistry , Hydrogen Bonding , Molecular Dynamics Simulation , Monosaccharide Transport Proteins/chemistry , Monosaccharide Transport Proteins/genetics , Mutation , Protein Binding , Protein Conformation , ThermodynamicsABSTRACT
Three shape-persistent [4+4] imine cages with truncated tetrahedral geometry with different window sizes were studied as hosts for the encapsulation of tetra-n-alkylammonium salts of various bulkiness. In various solvents the cages behave differently. For instance, in dichloromethane the cage with smallest window size takes up NEt4+ but not NMe4+, which is in contrast to the two cages with larger windows hosting both ions. To find out the reason for this, kinetic experiments were carried out to determine the velocity of uptake but also to deduce the activation barriers for these processes. To support the experimental results, calculations for the guest uptakes have been performed by molecular mechanics' simulations. Finally, the complexation of pharmaceutical interested compounds, such as acetylcholine, muscarine or denatonium have been determined by NMR experiments.
Subject(s)
Ammonium Compounds/chemistry , Imines/chemistry , Acetylcholine/chemistry , Density Functional Theory , Ions/chemistry , Kinetics , Magnetic Resonance Spectroscopy , Molecular Conformation , Muscarine/chemistry , Quaternary Ammonium Compounds/chemistry , Solvents/chemistry , ThermodynamicsABSTRACT
We designed and implemented a magnetic-driven scanning (MDS) probe for endoscopic optical coherence tomography (OCT). The probe uses an externally-driven tiny magnet in the distal end to achieve unobstructed 360-degree circumferential scanning at the side of the probe. The design simplifies the scanning part inside the probe and thus allows for easy miniaturization and cost reduction. We made a prototype probe with an outer diameter of 1.4 mm and demonstrated its capability by acquiring OCT images of ex vivo trachea and artery samples from a pigeon. We used a spectrometer-based Fourier-domain OCT system and the system sensitivity with our prototype probe was measured to be 91 dB with an illumination power of 850 µW and A-scan exposure time of 1 ms. The axial and lateral resolutions of the system are 6.5 µm and 8.1 µm, respectively.
