Bolhuis, Peter G.

Vreede, Jocelyne

Zhu, Lizhe

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Exploring Signal Transduction Mechanism of HAMP Domains via Molecular Dynamics and Metadynamics 

Monday, March 7, 2011 195aSwitch II has been identified as the main binding region between kinesin and
the microtubule surface.
1067-Plat
Molecular Dynamics Simulations of Monomeric and Dimeric NAC in
Alpha-Synuclein at Various Temperatures
Hongyi Yang1, James C. Patterson2.
1UAB, Birmingham, AL, USA, 2UAB, BIRMINGHAM, AL, USA.
The aggregation of a-Synuclein (aS), a 140-residue presynaptic protein, is
thought to be the primary step in the pathology of Parkinson’s disease. Several
studies have suggested that residues 61-95 of aS, known as the NAC region,
may play a critical role in promoting the aggregation of the full-length aS.
Furthermore, the first eighteen residues of NAC (i.e., residue 61-79) seem to
be essential for the self-assembly of NAC. To better understand the dynamic
structures of these proteins and mechanism by which they aggregate, molecular
dynamics (MD) simulations have been performed on the NAC region of aS. In
our MD simulations, the conformational changes of NAC are more sensitive to
increased temperatures than the full-length aS. The initial monomeric NAC
model, containing a-helix through the entire sequence, was obtained from the
NMRminimized average structure ofaS (PDB ID= 1XQ8). The dimeric peptide
modelswere constructed throughdocking.Atomistic simulations for bothmono-
meric and dimeric NAC in explicit water were conducted for 50 ns at 300 K and
372 K using the CHARMM22/CMAP force field. In simulations with mono-
meric NAC, the secondary structure of residues 74-84 and 87-92 were largely
unchanged at both 300 K and 372 K, while residues 61-73 lost the initial helical
structure much faster at 372 K than at 300 K. In simulations with dimeric NAC,
a short region encompassing residues 64-71 appeared a higher propensity to form
b structure than other regions. Inter-chain b-sheet structure was observed in this
region at the beginning of the simulation at 372 K; however, this b-sheet was in-
terrupted when the backbone of one peptide chain folded up, largely due to the
electrostatic attraction between residue 61 (i.e., Glu) and 80 (i.e., Lys).
1068-Plat
How Biomolecules Influence Water Structure and Dynamics
Ana Vila Verde1,2, R. Kramer Campen1.
1FOM institute AMOLF, Amsterdam, Netherlands, 2University of Minho,
Department of Physics, Braga, Portugal.
Interactions between solutes and water impact both water structure and struc-
tural dynamics as well as solute properties (e.g. conformational fluctuations
of proteins). To understand these interactions we investigate water near disac-
charides using classical atomistic molecular dynamics simulations. Disaccha-
rides show topological and chemical complexity characteristic of larger
biomolecules but are sufficiently small to permit detailed study. We observe
that increases in hydrophobicity precisely map slow down in water translation
and rotation of local water populations. In line with recent studies of proteins,
we find that chemically similar functional groups may interact differently with
water depending on neighboring functional groups.
To explain these observations we examine the mechanism of hydrogen bond
exchange for waters hydrogen bonded to other waters but within the sugar first
solvation shell, as well as waters hydrogen bonded to the sugar. Recent work
showed that water in bulk rotates through large angular jumps that pass through
bifurcated hydrogen bond intermediates and that rotation rates can be rational-
ized through transition state theory. Previous reports found that the rotational
slow down of water near small solutes can be predicted from changes in the ac-
cessible transition state volume or the enthalpy of the hydrogen bonds. For our
larger solutes we find that accounting for the transition state volume alone over-
estimates water rotational slow down. Differences in hydrogen bond enthalpy
are also insufficient to predict rotational slowdown. Water slowdown can
only be understood by additionally accounting for subtle changes in the free en-
ergy landscape associated with water rotation - reduction in the number of
available reactant states and broadening of the transition state barriers. The
presence of solutes of even moderate size thus affects water dynamics in
ways difficult to predict using simple scaling considerations from bulk, making
water response system dependent.
1069-Plat
Exploring Signal Transduction Mechanism of HAMP Domains via Molec-
ular Dynamics and Metadynamics
Lizhe Zhu, Peter G. Bolhuis, Jocelyne Vreede.
University of Amsterdam, Amsterdam, Netherlands.
The HAMP domain is a linker region in prokaryotic sensor proteins which
functions in two-component signal transduction pathways. HAMP exhibits
a parallel coiled coil motif comprising four helices and transfers the signal
from the sensor domain to the transmitter domain. We perfomed MD simula-
tions of a number of HAMP structures in isolation. These structures include
wild type and two mutants(A291V, A291F) of HAMP from A. fulgidus(PDB2ASW) and three HAMPs in the triHAMP chain (PDB 3LNR) from the P. aer-
uginosa soluble receptor Aer2. Our simulations show that a) 2ASW retains its
NMR structure in isolation; b) mutants of 2ASW exhibit slight higher flexibility
but relax fast to NMR 2ASW if mutations are removed; c) while the N-terminal
HAMP structure in the triHAMP chain is stable when isolated, the C-terminal
domain and the inner HAMP show extensive rearrangements, indicating that
the N-terminal constrains the flexibility of the triHAMP complex. Using MD
in combination with adaptive biasing potentials, known as the metadynamics
method, we explored different features of the signal transduction mechanism
of HAMP. Our results indicate that HAMP can transfer signals by changing
the tilt angle, the hydrophobic packing in the core of the complex and the ro-
tation angle of the helices. The implications of these results for signal transduc-
tion proteins containing HAMP are discussed.
1070-Plat
Dominant Site Involved in Inhibition of Targets by Anesthetics Suggested
Through Novel Free Energy Perturbation Calculations
David LeBard1, Grace Brannigan1, Jerome Henin2, Roderic Eckenhoff3,
Michael Klein1.
1Temple University, Philadelphia, PA, USA, 2CNRS - Marseille, Marseille,
France, 3University of Pennsylvania, Philadelphia, PA, USA.
It is well-established that general anesthetics bind directly to many of their tar-
gets, but the location of binding sites in a well-investigated target family, the
cys-loop receptors, remains unresolved. Although the structure of eukaryotic
cys-loop receptors has proven challenging to determine crystographically, a re-
cent prokaryotic anesthetic-sensitive member of the family (GLIC) has been
crystallized in complex with both a volatile and injected anesthetic. Detergent
molecules occupy the pore of both the apo and ligandedGLIC receptor, prevent-
ing any direct observation of pore-block by anesthetics from the crystal structure.
In recent microsecond atomisticmolecular dynamics (MD) simulations of flood-
ing by isoflurane of both the eukaryotic nicotinic acetylcholine receptor (nAChR)
and GLIC, we observed two isoflurane molecules bind to the pore of each recep-
tor. Circumventing the experimental obstacles, computational measurements of
binding affinity to the pore site provide an alternate method for determining the
relevance of pore-block to GLIC inhibition by anesthetics. In addition to a calcu-
lated binding affinity for isoflurane in the pore that correspondswell to themicro-
molar concentrations at which inhibition is observed, we also present novel
measurements using the alchemical free energy perturbation method (FEP) to
quantify negative cooperativity between the two isoflurane molecules in the
pore. The low Hill coefficient (&lt;0.5) calculated in this manner corresponds
to that measured experimentally within 10%. The results suggest that the pore
is probably occupied in GLIC exposed to experimental concentrations of isoflur-
ane, and pore-block may provide the dominant source of inhibition.
1071-Plat
Computer Simulation Studies of an Antimicrobial Peptide and aMicrobial
Virulence Factor: Statistical Convergence of Equilibrium Properties from
Massive Sampling
Chris Neale1,2, Re´gis Pome`s1,2.
1Department of Biochemistry, University of Toronto, Toronto, ON, Canada,
2Molecular Structure and Function, The Hospital for Sick Children, Toronto,
ON, Canada.
Critical to the immune system of eukaryotes, antimicrobial peptides (AMPs)
provide a line of defence against invading bacteria, often disrupting the integ-
rity of the bacterial membrane. Concurrently, bacteria repair this damage by the
action of their own integral membrane proteins. To gain insight into the molec-
ular basis of AMP action and membrane repair, we used one third of the world’s
28th fastest computer for one year to run massively repeated molecular dynam-
ics simulations of one such antagonistic pair: the eukaryotic AMP indolicidin
and the bacterial outer membrane enzyme and virulence factor PagP in explicit
membranes. Attaining adequate statistical sampling of conformational space
from molecular simulations of peptides and proteins in membranes is challeng-
ing because of the long correlation times of lipid bilayers. We analyze the con-
vergence of equilibrium properties in these two systems. We first investigate
the binding free energy of indolicidin to a lipid bilayer using umbrella sam-
pling, simulating each window for 1.5 microseconds and repeating this simula-
tion 30 times using different starting conformations. These simulations
demonstrate the existence of long-lived metastable states, particularly when
the peptide resides at or near the lipid-water interface. We show how many in-
dependent simulations reveal information that would likely be missed or mis-
interpreted if only a single simulation was performed. Second, we characterize
the functional binding mode of PagP, a bacterial outer membrane acyltransfer-
ase that binds and catalyses its lipid solvent. 100 independent 5-microsecond
simulations reveal the functional binding mode and associated gating mecha-
nism. From these simulations, which total 2 milliseconds of simulation time,
it is clear that the difficulty of reaching statistical convergence in atomistic


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Exploring Signal Transduction Mechanism of HAMP Domains via Molecular Dynamics and Metadynamics

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