Abstract
Using molecular dynamics (MD) simulation methods, we attempted to explain the experimental results on ligand specificity of glucose/galactose-binding protein (GGBP) to β-D-glucose and β-D-galactose. For the simulation, a three-dimensional structure of GGBP was prepared, and homology modeling was performed to generate variant structures of GGBP with mutations at Asp14. Then, docking was carried out to find a reasonable β-D-glucose and β-D-galactose binding conformations with GGBP. Subsequent molecular dynamics simulations of β-D-glucose–GGBP and β-D-galactose–GGBP complexes and estimation of the orientation and stability of water molecules at the binding site revealed how water molecules influence ligand specificity. In our simulation, water molecules mediated interactions of β-D-glucose or β-D-galactose with residue 14 of GGBP. In this mechanism, the Phe16Ala mutant leaves both sugar molecules free to move, and the specific role of water molecules were eliminated, while the wild type, Asp14Asn mutant, and Asp14Glu mutant make hydrogen bond interactions with β-D-glucose more favorable. Our results demonstrate that bound water molecules at the binding site of GGBP are related to localized conformational change, contributing to ligand specificity of GGBP for β-D-glucose over β-D-galactose.
Highlights
Periplasmic binding proteins (PBPs) exist in the periplasmic space between the inner membrane and outer membrane of gram-negative bacteria[1]
The results showed that the weakness of the hydrogen bond between the fourth hydroxyl group of galactose and Asp[14] was induced by localized conformational changes of Asp[14]
An molecular dynamics (MD) simulation was employed here to describe atomic fluctuations of sugar– Glucose/galactose-binding protein (GGBP) complexes, and GGBPs showed more favorable binding energies for glucose; this finding was consistent with experimental data on the binding affinity
Summary
Periplasmic binding proteins (PBPs) exist in the periplasmic space between the inner membrane and outer membrane of gram-negative bacteria[1]. In X-ray crystallographic studies, the PBP family has been shown to react rapidly with its ligand substrates with high affinity[2,3] Because of this property, PBPs are highly valued for industrial use as components of biosensor systems involving a fluorescent nanosensor[4,5,6]. In order to use GGBP as a glucose-monitoring biosensor, this protein should have strict sugar selectivity for glucose with high binding affinity. Despite the minor difference between glucose and galactose, wild-type GGBP shows distinct stereoselectivity for each sugar. To turn GGBP into a glucose sensor, many studies based on protein-engineering have been conducted[7,8,13], and one of the studies has shown that Asp14-mutated GGBPs (as a result of site-directed mutagenesis) have glucose-selective binding affinity under controlled experimental conditions[14].
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