bi6b00295_si_002.avi (12.36 MB)
Structural Determinants of Improved Fluorescence in a Family of Bacteriophytochrome-Based Infrared Fluorescent Proteins: Insights from Continuum Electrostatic Calculations and Molecular Dynamics Simulations
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posted on 2016-07-29, 20:19 authored by Mikolaj Feliks, Céline Lafaye, Xiaokun Shu, Antoine Royant, Martin FieldUsing
X-ray crystallography, continuum electrostatic calculations,
and molecular dynamics simulations, we have studied the structure,
protonation behavior, and dynamics of the biliverdin chromophore and
its molecular environment in a series of genetically engineered infrared
fluorescent proteins (IFPs) based on the chromophore-binding domain
of the Deinococcus radiodurans bacteriophytochrome.
Our study suggests that the experimentally observed enhancement of
fluorescent properties results from the improved rigidity and planarity
of the biliverdin chromophore, in particular of the first two pyrrole
rings neighboring the covalent linkage to the protein. We propose
that the increases in the levels of both motion and bending of the
chromophore out of planarity favor the decrease in fluorescence. The
chromophore-binding pocket in some of the studied proteins, in particular
the weakly fluorescent parent protein, is shown to be readily accessible
to water molecules from the solvent. These waters entering the chromophore
region form hydrogen bond networks that affect the otherwise planar
conformation of the first three rings of the chromophore. On the basis
of our simulations, the enhancement of fluorescence in IFPs can be
achieved either by reducing the mobility of water molecules in the
vicinity of the chromophore or by limiting the interactions of the
nearby protein residues with the chromophore. Finally, simulations
performed at both low and neutral pH values highlight differences
in the dynamics of the chromophore and shed light on the mechanism
of fluorescence loss at low pH.