Kin Lam and Emad Tajkhorshid.
Membrane interactions of Cy3/Cy5 fluorophores and their effects on
membrane protein dynamics.
Biophysical Journal, 119:24-34, 2020.
(PMC: PMC7335937)
LAM2020-ET
Organic fluorophores, such as Cy3 and Cy5, have been widely used as
chemical labels to probe
the structure and dynamics of membrane proteins. While a number of
previous studies have reported
on the possibility of some of the water-soluble fluorophores to
interact with lipid bilayers,
detailed fluorophore-lipid interactions and, more importantly, the
potential effect of such interactions
on the natural dynamics of the labeled membrane proteins have not been
well studied.
We have performed a large set of all-atom molecular dynamics
simulations employing the
highly mobile membrane mimetic model to describe spontaneous
partitioning of
the fluorophores into lipid bilayers with different lipid compositions.
Spontaneous membrane partitioning of Cy3 and Cy5 fluorophores captured
in
these simulations proceeds in two steps.
Electrostatic interaction between the fluorophores and the lipid
headgroups facilitates the initial, fast
membrane association of the fluorophores, followed by slow insertion of
hydrophobic moieties
into the lipid bilayer core.
After the conversion of the resulting membrane-bound systems to full-
membrane representations,
biased-exchange umbrella sampling simulations are performed for free
energy calculations,
revealing a higher energy barrier for partitioning into negatively
charged (PS/PC)
membranes than purely zwitterionic (PC or PE) ones. Furthermore, the
potential effect of
fluorophore-lipid interactions
on membrane proteins has been examined by covalently linking Cy5 to
single- and
multi-pass transmembrane helical proteins.
Equilibrium simulations show strong position-dependent effects of Cy5-
tagging
on the structure and natural dynamics of membrane proteins.
Interactions between the tagged protein and Cy5 were also observed.
Our results suggest that fluorophore-lipid interactions
can affect the structure and dynamics of membrane proteins to various
extents, specially in systems
with higher structural flexibility.
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