10 research outputs found
Origin Of Fluorescence In 11-cis Locked Bovine Rhodopsin
The excited state lifetime of bovine rhodopsin (Rh) increases from ca. 100 fs to 85 ps when the C11=C12 bond of its chromophore is locked by a cyclopentene moiety (Rh5). To explain such an increase, we employed ab initio multiconfigurational quantum chemistry to construct computer models of Rh and Rh5 and to investigate the shape of their excited state potential energy surfaces in a comparative way. Our results show that the observed Rh5 fluorescence (lambda(f)(max) = 620 nm) is due to a previously unreported locally excited intermediate whose lifetime is controlled by a small energy barrier. The analysis of the properties and decay path of such an intermediate provides useful information for engineering rhodopsin variants with augmented fluorescence efficiencies
THE FLUORESCENCE OF THE WURSTER'S BLUE RADICAL CATION IS CONTROLLED BY A CONICAL INTERSECTION
Author Institution: Department of Chemistry and Center for Photochemical Sciences, Bowling Green State University, Bowling Green, OH 43403, USA; Physical Chemistry Department, University of Geneva, 1211 Geneva, Switzerland\indent The photochemistry and photophysics of a stable N,N,N',N'- tetramethyl-\textit{p}-phenylenediamine radical cation (commonly known as Wurster s Blue) is the subject of current research interest as it represents an example of mixed valence (MV) compound. In this work we used \textit{ab initio} CASSCF/CASPT2 quantum chemical calculations to map its first excited state (D) potential energy surface in the gas-phase. \indent According to the spectral data by Grilj et al., the fluorescence of Wurster s Blue radical cation could only be observed at low temperatures (below 115K). In order to explain this behavior, the conical intersection space (IS) between the first excited (D) and the ground state (D) potential energy surfaces was mapped and characterized. The intrinsic reaction coordinate (IRC) scan, following the relaxation of the Wurster s blue molecule from the D/D intersection space along the D potential energy surface, led to the ground state equilibrium structure. The energy barrier between the excited state energy minimum and the lowest lying conical intersection structure (CI) was calculated to be 3.1 kcal/mol. As a result, we concluded that this barrier was responsible for the observed temperature dependence of the fluorescence that disappears at temperatures above 115K due to the opening of a radiationless deactivation channel
Origin of Fluorescence in 11-cisLocked Bovine Rhodopsin
The excited state lifetime of bovine rhodopsin (Rh) increases from ca. 100 fs to 85 ps when the C11=C12 bond of its chromophore is locked by a cyclopentene moiety (Rh5). To explain such an increase, we employed ab initio multiconfigurational quantum chemistry to construct computer models of Rh and Rh5 and to investigate the shape of their excited state potential energy surfaces in a comparative way. Our results show that the observed Rh5 fluorescence (lambda(f)(max) = 620 nm) is due to a previously unreported locally excited intermediate whose lifetime is controlled by a small energy barrier. The analysis of the properties and decay path of such an intermediate provides useful information for engineering rhodopsin variants with augmented fluorescence efficiencies
pH-Dependent Transient Conformational States Control Optical Properties in Cyan Fluorescent Protein
A recently engineered mutant of cyan
fluorescent protein (WasCFP)
that exhibits pH-dependent absorption suggests that its tryptophan-based
chromophore switches between neutral (protonated) and charged (deprotonated)
states depending on external pH. At pH 8.1, the latter gives rise
to green fluorescence as opposed to the cyan color of emission that
is characteristic for the neutral form at low pH. Given the high energy
cost of deprotonating the tryptophan at the indole nitrogen, this
behavior is puzzling, even if the stabilizing effect of the V61K mutation
in proximity to the protonation/deprotonation site is considered.
Because of its potential to open new avenues for the development of
optical sensors and photoconvertible fluorescent proteins, a mechanistic
understanding of how the charged state in WasCFP can possibly be stabilized
is thus important. Attributed to the dynamic nature of proteins, such
understanding often requires knowledge of the various conformations
adopted, including transiently populated conformational states. Transient
conformational states triggered by pH are of emerging interest and
have been shown to be important whenever ionizable groups interact
with hydrophobic environments. Using a combination of the weighted-ensemble
sampling method and explicit-solvent constant pH molecular dynamics
(CPHMD<sup>MSλD</sup>) simulations, we have identified a solvated
transient state, characterized by a partially open β-barrel
where the chromophore p<i>K</i><sub>a</sub> of 6.8 is shifted
by over 20 units from that of the closed form (6.8 and 31.7, respectively).
This state contributes a small population at low pH (12% at pH 6.1)
but becomes dominant at mildly basic conditions, contributing as much
as 53% at pH 8.1. This pH-dependent population shift between neutral
(at pH 6.1) and charged (at pH 8.1) forms is thus responsible for
the observed absorption behavior of WasCFP. Our findings demonstrate
the conditions necessary to stabilize the charged state of the WasCFP
chromophore (namely, local solvation at the deprotonation site and
a partial flexibility of the protein β-barrel structure) and
provide the first evidence that transient conformational states can
control optical properties of fluorescent proteins
Deconstructing Activation Events in Rhodopsin
Activation of class-A
G-protein-coupled receptors (GPCRs) involves
large-scale reorganization of the H3/H6 interhelical network. In rhodopsin
(Rh), this process is coupled to a change in the protonation state
of a key residue, E134, whose exact role in activation is not well
understood. Capturing this millisecond pH-dependent process is a well-appreciated
challenge. We have developed a scheme combining the harmonic Fourier
beads (HFB) method and constant-pH molecular dynamics with pH-based
replica exchange (pH-REX) to gain insight into the structural changes
that occur along the activation pathway as a function of the protonation
state of E134. Our results indicate that E134 is protonated as a consequence
of tilting of H6 by ca. 4.0° with respect to its initial position
and simultaneous rotation by ca. 23° along its principal axis.
The movement of H6 is associated with breakage of the E247–R135
and R135–E134 salt bridges and concomitant release of the E134
side chain, which results in an increase in its p<i>K</i><sub>a</sub> value above physiological pH. An increase in the hydrophobicity
of the environment surrounding E134 leads to further tilting and rotation
of H6 and upshift of the E134 p<i>K</i><sub>a</sub>. Such
atomic-level information, which is not accessible through experiments,
refines the earlier proposed sequential model of Rh activation (see: Zaitseva,
E.; et al. Sequential Rearrangement
of Interhelical Networks Upon Rhodopsin Activation in Membranes: The
Meta II<sub>a</sub> Conformational Substate. J. Am. Chem. Soc. 2010, 132, 4815) and argues that the E134 protonation
switch is both a cause and a consequence of the H6 motion
Origin of Fluorescence in 11-<i>cis</i> Locked Bovine Rhodopsin
The excited state lifetime of bovine rhodopsin (Rh) increases
from
ca. 100 fs to 85 ps when the C11î—»C12 bond of its chromophore
is locked by a cyclopentene moiety (Rh5). To explain such an increase,
we employed <i>ab initio</i> multiconfigurational quantum
chemistry to construct computer models of Rh and Rh5 and to investigate
the shape of their excited state potential energy surfaces in a comparative
way. Our results show that the observed Rh5 fluorescence (λ<sub>max</sub><sup>f</sup> = 620 nm) is due to a previously unreported
locally excited intermediate whose lifetime is controlled by a small
energy barrier. The analysis of the properties and decay path of such
an intermediate provides useful information for engineering rhodopsin
variants with augmented fluorescence efficiencies