8 research outputs found
Specific Ion Interaction Dominates over Hydrophobic Matching Effects in PeptideāLipid Bilayer Interactions: The Case of Short Peptide
Insertion of short peptides into
the cell membrane is energetically
unfavorable and challenges the commonly accepted hydrophobic matching
principle. Yet there has been evidence that many short peptides can
penetrate into the cells to perform the biological functions in salt
solution. On the basis of the previous study (J. Phys. Chem. C 2013, 117, 11095ā11103), here we further performed a systematic study on the
interaction of mastoparan with various neutral lipid bilayers with
different lipid chain lengths in situ to examine the hydrophobic matching
principle in different aqueous salt environments using sum frequency
generation vibrational spectroscopy. It is found that the hydrophobic
matching is the dominant driving force for the association of MP with
a lipid bilayer in a pure water environment. However, in a kosmotropic
ion environment, the hydration of ions can overcome the hydrophobic
mismatching effects, leading to the insertion of MP into lipid bilayers
with much longer hydrophobic lengths. When the hydrophobic thickness
of the bilayer is much longer than MPās hydrophobic length,
MP diffuses on a single monolayer, rather than spanning the bilayer
to prevent the exposure of the hydrophilic part of MP to the lipid
hydrophobic moiety. Findings from the present study suggest that the
interaction between the positively charged choline group of a lipid
and kosmotropic ions could be an important step for effective peptide
insertion into a cell membrane. Results from our studies will provide
an insight into how the short peptides form the ion channel in a thick
membrane and offer some ideas for cellular delivery
Phosphate Ions Promoting Association between Peptide and Modeling Cell Membrane Revealed by Sum Frequency Generation Vibrational Spectroscopy
Phosphate
ion is one of the most important anions present in the intracellular
and extracellular fluid. It can form strongly hydrogen-bonded and
salt-bridged complexes with arginine and lysine to activate the voltage
gated channel protein. A molecular-level insight into how the phosphate
anions mediate the interaction between peptides and cell membrane
is critical to understand membrane-bound peptide actions. In this
study, sum frequency generation vibrational spectroscopy (SFG-VS)
has been applied to characterize interactions between mastoparan (MP,
a G-protein-activating peptide) and different charged lipid bilayers
in situ. It is found that phosphate ions can greatly promote the association
of MP with lipid bilayers and accelerate the conformation transition
of membrane-bound MP from aggregation into Ī±-helical structure.
In phosphate buffer solution, MP can insert not only into negatively
and neutrally charged lipid bilayers but also into positively charged
lipid bilayers. In neutrally and negatively charged lipid bilayers,
the tilt angle of Ī±-helical structure becomes smaller with increasing
buffer concentration, while MP adopts a multiple orientation distribution
in the positively charged lipid bilayer. MP interacts with lipid bilayers
in the salt solution environment most likely by formation of toroidal
pores inside the bilayer matrix. Results from our studies will provide
insight into the MP action mechanism and offer some ideas to deliver
exogenous protein into the cytosol
Accurate Determination of Interfacial Protein Secondary Structure by Combining Interfacial-Sensitive Amide I and Amide III Spectral Signals
Accurate determination of protein
structures at the interface is
essential to understand the nature of interfacial protein interactions,
but it can only be done with a few, very limited experimental methods.
Here, we demonstrate for the first time that sum frequency generation
vibrational spectroscopy can unambiguously differentiate the interfacial
protein secondary structures by combining surface-sensitive amide
I and amide III spectral signals. This combination offers a powerful
tool to directly distinguish random-coil (disordered) and Ī±-helical
structures in proteins. From a systematic study on the interactions
between several antimicrobial peptides (including LKĪ±14, mastoparan
X, cecropin P1, melittin, and pardaxin) and lipid bilayers, it is
found that the spectral profiles of the random-coil and Ī±-helical
structures are well separated in the amide III spectra, appearing
below and above 1260 cm<sup>ā1</sup>, respectively. For the
peptides with a straight backbone chain, the strength ratio for the
peaks of the random-coil and Ī±-helical structures shows a distinct
linear relationship with the fraction of the disordered structure
deduced from independent NMR experiments reported in the literature.
It is revealed that increasing the fraction of negatively charged
lipids can induce a conformational change of pardaxin from random-coil
to Ī±-helical structures. This experimental protocol can be employed
for determining the interfacial protein secondary structures and dynamics
in situ and in real time without extraneous labels
Molecular Binding Sites Are Located Near the Interface of Intrinsic Dynamics Domains (IDDs)
We provide evidence supporting that
proteināprotein and
proteināligand docking poses are functions of protein shape
and intrinsic dynamics. Over sets of 68 proteināprotein complexes
and 240 nonhomologous enzymes, we recognize common predispositions
for binding sites to have minimal vibrations and angular momenta,
while two interacting proteins orient so as to maximize the angle
between their rotation/bending axes (>65Ā°). The findings are
then used to define quantitative criteria to filter out docking decoys
less likely to be the near-native poses; hence, the chances to find
near-native hits can be doubled. With the novel approach to partition
a protein into ādomainsā of robust but disparate intrinsic
dynamics, 90% of catalytic residues in enzymes can be found within
the first 50% of the residues closest to the interface of these dynamics
domains. The results suggest an anisotropic rather than isotropic
distribution of catalytic residues near the mass centers of enzymes
In Situ Molecular-Level Insights into the Interfacial Structure Changes of Membrane-Associated Prion Protein Fragment [118ā135] Investigated by Sum Frequency Generation Vibrational Spectroscopy
Protein aggregation is associated with many āprotein
deposition
diseasesā. A precise molecular detail of the conformational
transitions of such a membrane-associated protein structure is critical
to understand the disease mechanism and develop effective treatments.
One potential model peptide for studying the mechanism of protein
deposition diseases is prion protein fragment [118ā135] (PrP118ā135),
which shares homology with the C-terminal domain of the Alzheimerās
Ī²-amyloid peptide. In this study, sum frequency generation vibrational
spectroscopy (SFG-VS) has been applied to characterize interactions
between PrP118ā135 and 1-palmitoyl-2-oleoyl-<i>sn</i>-glycero-3-phospho-(1ā²-<i>rac</i>-glycerol) (POPG)
lipid bilayer in situ. The conformation change and orientation of
PrP118ā135 in lipid bilayers have been determined using SFG
spectra with different polarization combinations. It is found that
low-concentration PrP118ā135 predominantly adopts Ī±-helical
structure but with tiny Ī²-sheet structure. With the PrP118ā135
concentration increasing, the molecular number ratio of parallel Ī²-sheet
structure increases and reaches about 44% at a concentration of 0.10 mg/mL,
indicating the formation of abnormally folded scrapie isoforms. The
Ī±-helical structure inserts into the lipid bilayer with a tilt
angle of ā¼32Ā° versus the surface normal, while the Ī²-sheet
structure lies down on the lipid bilayer with the tilt and twist angle
both of 90Ā°. The 3300 cm<sup>ā1</sup> NāH stretching
signal in psp spectra arises from Ī±-helical structure at low
PrP concentration and from the Ī²-sheet structure at high PrP
concentration. Results from this study will provide an in-depth insight
into the early events in the aggregation of PrP in cell membrane
Effect of Dehydration on the Interfacial Water Structure at a Charged Polymer Surface: Negligible Ļ<sup>(3)</sup> Contribution to Sum Frequency Generation Signal
Interfacial water structure at charged surfaces plays
a key role
in many physical, chemical, biological, environmental, and industrial
processes. Understanding the release of interfacial water from the
charged solid surfaces during dehydration process may provide insights
into the mechanism of protein folding and the nature of weak molecular
interactions. In this work, sum frequency generation vibrational spectroscopy
(SFG-VS), supplemented by quartz crystal microbalance (QCM) measurements,
has been applied to study the interfacial water structure at polyelectrolyte
covered surfaces. PolyĀ[2-(dimethylamino)Āethyl methacrylate] (PDMAEMA)
chains are grafted on solid surfaces to investigate the change of
interfacial water structure with varying surface charge density induced
by tuning the solution pH. At pH ā¤ 7.1, SFG-VS intensity is
linear to the loss of mass of interfacial water caused by the dehydration
of PDMAEMA chains, and no reorientation of the strongly bonded water
molecules is observed in the light of Ļ<sub>ppp</sub>/Ļ<sub>ssp</sub> ratio. Ļ<sup>(3)</sup> contribution to SFG signal
is deduced based on the combination of SFG and QCM results. It is
the first direct experimental evidence to reveal that the Ļ<sup>(3)</sup> has a negligible contribution to SFG signal of the interfacial
water at a charged polymer surface
In Situ and Real-Time SFG Measurements Revealing Organization and Transport of Cholesterol Analogue 6āKetocholestanol in a Cell Membrane
Cholesterol
organization and transport within a cell membrane are
essential for human health and many cellular functions yet remain
elusive so far. Using cholesterol analogue 6-ketocholestanol (6-KC)
as a model, we have successfully exploited sum frequency generation
vibrational spectroscopy (SFG-VS) to track the organization and transport
of cholesterol in a membrane by combining achiral-sensitive ssp (ppp)
and chiral-sensitive psp polarization measurements. It is found that
6-KC molecules are aligned at the outer leaflet of the DMPC lipid
bilayer with a tilt angle of about 10Ā°. 6-KC organizes itself
by forming an Ī±āĪ² structure at low 6-KC concentration
and most likely a Ī²āĪ² structure at high 6-KC concentration.
Among all proposed models, our results favor the so-called umbrella
model with formation of a 6-KC cluster. Moreover, we have found that
the long anticipated flip-flop motion of 6-KC in the membrane takes
time to occur, at least much longer than previously thought. All of
these interesting findings indicate that it is critical to explore
in situ, real-time, and label-free methodologies to obtain a precise
molecular description of cholesterolās behavior in membranes.
This study represents the first application of SFG to reveal the cholesterolālipid
interaction mechanism at the molecular level
Observing a Model Ion Channel Gating Action in Model Cell Membranes in Real Time in Situ: Membrane Potential Change Induced Alamethicin Orientation Change
Ion channels play crucial roles in transport and regulatory
functions
of living cells. Understanding the gating mechanisms of these channels
is important to understanding and treating diseases that have been
linked to ion channels. One potential model peptide for studying the
mechanism of ion channel gating is alamethicin, which adopts a split
Ī±/3<sub>10</sub>-helix structure and responds to changes in
electric potential. In this study, sum frequency generation vibrational
spectroscopy (SFG-VS), supplemented by attenuated total reflectance
Fourier transform infrared spectroscopy (ATR-FTIR), has been applied
to characterize interactions between alamethicin (a model for larger
channel proteins) and 1-palmitoyl-2-oleoyl-<i>sn</i>-glycero-3-phosphocholine
(POPC) lipid bilayers in the presence of an electric potential across
the membrane. The membrane potential difference was controlled by
changing the pH of the solution in contact with the bilayer and was
measured using fluorescence spectroscopy. The orientation angle of
alamethicin in POPC lipid bilayers was then determined at different
pH values using polarized SFG amide I spectra. Assuming that all molecules
adopt the same orientation (a Ī“ distribution), at pH = 6.7 the
Ī±-helix at the N-terminus and the 3<sub>10</sub>-helix at the
C-terminus tilt at about 72Ā° (Īø<sub>1</sub>) and 50Ā°
(Īø<sub>2</sub>) versus the surface normal, respectively. When
pH increases to 11.9, Īø<sub>1</sub> and Īø<sub>2</sub> decrease
to 56.5Ā° and 45Ā°, respectively. The Ī“ distribution
assumption was verified using a combination of SFG and ATR-FTIR measurements,
which showed a quite narrow distribution in the angle of Īø<sub>1</sub> for both pH conditions. This indicates that all alamethicin
molecules at the surface adopt a nearly identical orientation in POPC
lipid bilayers. The localized pH change in proximity to the bilayer
modulates the membrane potential and thus induces a decrease in both
the tilt and the bend angles of the two helices in alamethicin. This
is the first reported application of SFG to the study of model ion
channel gating mechanisms in model cell membranes