45 research outputs found
Mechanism of lignin inhibition of enzymatic biomass deconstruction
Background
The conversion of plant biomass to ethanol via enzymatic cellulose hydrolysis offers a potentially sustainable route to biofuel production. However, the inhibition of enzymatic activity in pretreated biomass by lignin severely limits the efficiency of this process. Results
By performing atomic-detail molecular dynamics simulation of a biomass model containing cellulose, lignin, and cellulases (TrCel7A), we elucidate detailed lignin inhibition mechanisms. We find that lignin binds preferentially both to the elements of cellulose to which the cellulases also preferentially bind (the hydrophobic faces) and also to the specific residues on the cellulose-binding module of the cellulase that are critical for cellulose binding of TrCel7A (Y466, Y492, and Y493). Conclusions Lignin thus binds exactly where for industrial purposes it is least desired, providing a simple explanation of why hydrolysis yields increase with lignin removal
ChAdOx1 interacts with CAR and PF4 with implications for thrombosis with thrombocytopenia syndrome
Vaccines derived from chimpanzee adenovirus Y25 (ChAdOx1), human adenovirus type 26 (HAdV-D26), and human adenovirus type 5 (HAdV-C5) are critical in combatting the severe acute respiratory coronavirus 2 (SARS-CoV-2) pandemic. As part of the largest vaccination campaign in history, ultrarare side effects not seen in phase 3 trials, including thrombosis with thrombocytopenia syndrome (TTS), a rare condition resembling heparin-induced thrombocytopenia (HIT), have been observed. This study demonstrates that all three adenoviruses deployed as vaccination vectors versus SARS-CoV-2 bind to platelet factor 4 (PF4), a protein implicated in the pathogenesis of HIT. We have determined the structure of the ChAdOx1 viral vector and used it in state-of-the-art computational simulations to demonstrate an electrostatic interaction mechanism with PF4, which was confirmed experimentally by surface plasmon resonance. These data confirm that PF4 is capable of forming stable complexes with clinically relevant adenoviruses, an important step in unraveling the mechanisms underlying TTS.
Abstract
INTRODUCTION
RESULTS
DISCUSSION
MATERIALS AND METHODS
Acknowledgments
Supplementary Materials
REFERENCES AND NOTES
0eLetters
Abstract
Vaccines derived from chimpanzee adenovirus Y25 (ChAdOx1), human adenovirus type 26 (HAdV-D26), and human adenovirus type 5 (HAdV-C5) are critical in combatting the severe acute respiratory coronavirus 2 (SARS-CoV-2) pandemic. As part of the largest vaccination campaign in history, ultrarare side effects not seen in phase 3 trials, including thrombosis with thrombocytopenia syndrome (TTS), a rare condition resembling heparin-induced thrombocytopenia (HIT), have been observed. This study demonstrates that all three adenoviruses deployed as vaccination vectors versus SARS-CoV-2 bind to platelet factor 4 (PF4), a protein implicated in the pathogenesis of HIT. We have determined the structure of the ChAdOx1 viral vector and used it in state-of-the-art computational simulations to demonstrate an electrostatic interaction mechanism with PF4, which was confirmed experimentally by surface plasmon resonance. These data confirm that PF4 is capable of forming stable complexes with clinically relevant adenoviruses, an important step in unraveling the mechanisms underlying TTS
Atomistic Origins of Biomass Recalcitrance in Organosolv Pretreatment
Secondary plant cell walls are made from three common biopolymers, cellulose, lignin, and hemicellulose, representing a critical feedstock for sustainable biomaterial production. Separating lignocellulosic biomass components for use in tailored sustainable energy and materials applications is challenging, as the biopolymers are in close proximity within the plant secondary cell wall. Organic solvents are used to pretreat recalcitrant biomass and separate the interacting polymers, solubilizing the lignin fraction for lignin-first valorization approaches. However, no single organosolv pretreatment approach has proven superior for heterogeneous biomass samples. Simulation offers a complementary atomic view into interactions between biomass components, resolving mechanistic hypotheses for how biomass composition influences separations processes. Using molecular dynamics simulations, we quantify lignin-cellulose interactions through binding free energies determined from 300 lignin polymer models in nine solvent environments, across four crystalline cellulose faces, with an aggregate simulation time of nearly 154 microseconds. The binding free energy determined from simulation categorizes the solvents. For poor lignin solvents, all lignin polymers bind strongly to cellulose. By contrast, polar protic solvents such as methanol and ethanol favor the unbinding between lignin and cellulose in all conditions, regardless of charge for the lignin monomer tested. Aprotic organic solvents separate lignin from cellulose only for uncharged lignin monomers, with charged lignin monomers associating to cellulose. While polar protic solvents are most effective at breaking apart lignin-cellulose interactions for charged lignin species, solvent dynamics highlight that there is no single optimal solvent to facilitate lignin-cellulose separation, particularly as some solvents demonstrate greater effectiveness for skewed S:G ratios. Instead, the optimal solvent for a given lignin sample will depend on the lignin compound and the net charge for the lignin polymers
Memory, gender and anti-fascism in France and Britain in the 1930s
Synaptotagmin
(Syt) is a membrane-associated protein involved in
vesicle fusion through the SNARE complex that is found throughout
the human body in 17 different isoforms. These isoforms have two membrane-binding
C2 domains, which sense Ca<sup>2+</sup> and thereby promote anionic
membrane binding and lead to vesicle fusion. Through molecular dynamics
simulations using the highly mobile membrane mimetic acclerated bilayer
model, we have investigated how small protein sequence changes in
the Ca<sup>2+</sup>-binding loops of the C2 domains may give rise
to the experimentally determined difference in binding kinetics between
Syt-1 and Syt-7 isoforms. Syt-7 C2 domains are found to form more
close contacts with anionic phospholipid headgroups, particularly
in loop 1, where an additional positive charge in Syt-7 draws the
loop closer to the membrane and causes the anchoring residue F167
to insert deeper into the bilayer than the corresponding methionine
in Syt-1 (M173). By performing additional replica exchange umbrella
sampling calculations, we demonstrate that these additional contacts
increase the energetic cost of unbinding the Syt-7 C2 domains from
the bilayer, causing them to unbind more slowly than their counterparts
in Syt-1
Reversible Unwrapping Algorithm for Constant-Pressure Molecular Dynamics Simulations
Molecular
simulation technologies have afforded researchers a unique
look into the nanoscale interactions driving physical processes. However,
a limitation for molecular dynamics (MD) simulations is that they
must be performed on finite-sized systems in order to map onto computational
resources. To minimize artifacts arising from finite-sized simulation
systems, it is common practice for MD simulations to be performed
with periodic boundary conditions (PBCs). However, in order to calculate
specific physical properties, such as mean square displacements to
calculate diffusion coefficients, continuous particle trajectories
where the atomic movements are continuous and do not jump between
cell faces are required. In these cases, modifying atomic coordinates
through unwrapping schemes is an essential post-processing tool to
remove these jumps. Here, two established trajectory unwrapping schemes
are applied to 1 μs wrapped trajectories for a small water box
and lysozyme in water. The existing schemes can result in spurious
diffusion coefficients, long bonds within unwrapped molecules, and
inconsistent atomic coordinates when coordinates are rewrapped after
unwrapping. We determine that prior unwrapping schemes do not account
for changing periodic box dimensions and introduce an additional correction
term to the existing displacement unwrapping scheme to correct for
these artifacts. We also demonstrate that the resulting algorithm
is a hybrid between the existing heuristic and displacement unwrapping
schemes. After treatment using this new unwrapping scheme, molecular
geometries are correct even after long simulations. In anticipation
for longer MD trajectories, we develop implementations for this new
scheme in multiple PBC handling tools
LongBondEliminator: A Molecular Simulation Tool to Remove Ring Penetrations in Biomolecular Simulation Systems
We develop a workflow, implemented as a plugin to the molecular visualization program VMD, that can fix ring penetrations with minimal user input. LongBondEliminator, detects ring piercing artifacts by the long, strained bonds that are the local minimum energy conformation during minimization for some assembled simulation system. The LongBondEliminator tool then automatically treats regions near these long bonds using multiple biases applied through NAMD. By combining biases implemented through the collective variables module, density-based forces, and alchemical techniques in NAMD, LongBondEliminator will iteratively alleviate long bonds found within molecular simulation systems. Through three concrete examples with increasing complexity, a lignin polymer, an viral capsid assembly, and a large, highly glycosylated protein aggrecan, we demonstrate the utility for this method in eliminating ring penetrations from classical MD simulation systems. The tool is available via gitlab as a VMD plugin, and has been developed to be generically useful across a variety of biomolecular simulations
A Microscopic View of Phospholipid Insertion into Biological Membranes
Understanding the process of membrane
insertion is an essential
step in developing a detailed mechanism, for example, for peripheral
membrane protein association and membrane fusion. The highly mobile
membrane mimetic (HMMM) has been used to accelerate the membrane association
and binding of peripheral membrane proteins in simulations by increasing
the lateral diffusion of phospholipid headgroups while retaining an
atomistic description of the interface. Through a comparative study,
we assess the difference in insertion rate of a free phospholipid
into an HMMM as well as into a conventional phospholipid bilayer and
develop a detailed mechanistic model of free phospholipid insertion
into biological membranes. The mechanistic insertion model shows that
successful irreversible association of the free phospholipid to the
membrane interface, which results in its insertion, is the rate-limiting
step. Association is followed by independent, sequential insertion
of the acyl tails of the free phospholipid into the membrane, with
splayed acyl tail intermediates. Use of the HMMM is found to replicate
the same intermediate insertion states as in the full phospholipid
bilayer; however, it accelerates overall insertion by approximately
a factor of 3, with the probability of successful association of phospholipid
to the membrane being significantly enhanced
Passive permeability controls synthesis for the allelochemical sorgoleone in sorghum root exudate
Competition for soil nutrients and water with other plants foster competition within the biosphere for access to these limited resources. The roots for the common grain sorghum produce multiple small molecules that are released via root exudates into the soil to compete with other plants. Sorgoleone is one such compound, which suppresses weed growth near sorghum by acting as a quinone analog and interferes with photosynthesis. Since sorghum also grows photosynthetically, and may be susceptible to sorgoleone action if present in tissues above ground, it is essential for sorgoleone to be excreted efficiently. However, since the P450 enzymes that synthesize sorgoleone are intracellular, the release mechanism for sorgoleone remain unclear. In this study, we conducted an in silico assessment for sorgoleone and its precursors to passively permeate biological membranes. To facilitate accurate simulation, CHARMM parameters were newly optimized for sorgoleone and its precursors. These parameters were used to conduct one microsecond of unbiased molecular dynamics simulations to compare the permeability of sorgoleone with its precursors molecules. We find that interleaflet transfer is maximized for sorgoleone, suggesting that the precursor molecules may remain in the same leaflet for access by biosynthetic P450 enzymes. Since no sorgoleone was extracted during unbiased simulations, we compute a permeability coefficient using the inhomogeneous solubility diffusion model. The requisite free energy and diffusivity profiles for sorgleone through a sorghum plasma membrane model were determined through Replica Exchange Umbrella Sampling (REUS) simulations. The REUS calculations highlight that any soluble sorgleone would quickly insert into a lipid bilayer, and would readily transit. When sorgleone forms aggregates in root exudate as indicated by our equilibrium simulations, aggregate formation would lower the effective concentration in aqueous solution, creating a concentration gradient that would facilitate passive transport. This suggests that sorgoleone synthesis occurs within sorghum root cells and that sorgoleone is exuded by permeating through the cell membrane without the need for a transport protein
Plant Terpenoid Permeability through Biological Membranes Explored via Molecular Simulations
Plants synthesize small molecule diterpenes comprised of twenty carbon from precursors isopentenyl diphosphate and dimethylallyl disphosphate, manufacturing diverse compounds used for defense, signaling, and other functions. Industrially, diterpenes are used as natural aromas and flavoring, as pharmaceuticals, and as natural insecticides or repellents. Despite diterpene ubiquity in plant systems, it remains unknown how plants control diterpene localization and transport. For many other small molecules, plant cells maintain transport proteins that control compound compartmentalization. However, for most diterpene compounds, specific transport proteins have not been identified, and so it has been hypothesized that diterpene may cross biological membranes passively. Through molecular simulation, we study membrane transport for three complex diterpenes from among the many made by members of the Lamiaceae family to determine their permeability coefficient across plasma membrane models. To facilitate accurate simulation, the intermolecular interactions for leubethanol, abietic acid, and sclareol were parameterized through the standard CHARMM methodology for incorporation into molecular simulations. To evaluate the effect of membrane composition on permeability, we simulate the three diterpenes in two membrane models derived from sorghum and yeast lipidomics data. We track permeation events within our unbiased simulations, and compare implied permeation coefficients with those calculated from Replica Exchange Umbrella Sampling calculations using the inhomogeneous solubility diffusion model. The diterpenes are observed to permeate freely through these membranes, indicating that a transport protein may not be needed to export these small molecules from plant cells. Moreover, the permeability is observed to be greater for plant-like membrane compositions when compared against animal-like membrane models. Increased permeability for diterpene molecules in plant membranes suggest that plants have tailored their membranes to facilitate low-energy transport processes for signaling molecules
Differential Membrane Binding Mechanics of Synaptotagmin Isoforms Observed in Atomic Detail
Synaptotagmin
(Syt) is a membrane-associated protein involved in
vesicle fusion through the SNARE complex that is found throughout
the human body in 17 different isoforms. These isoforms have two membrane-binding
C2 domains, which sense Ca<sup>2+</sup> and thereby promote anionic
membrane binding and lead to vesicle fusion. Through molecular dynamics
simulations using the highly mobile membrane mimetic acclerated bilayer
model, we have investigated how small protein sequence changes in
the Ca<sup>2+</sup>-binding loops of the C2 domains may give rise
to the experimentally determined difference in binding kinetics between
Syt-1 and Syt-7 isoforms. Syt-7 C2 domains are found to form more
close contacts with anionic phospholipid headgroups, particularly
in loop 1, where an additional positive charge in Syt-7 draws the
loop closer to the membrane and causes the anchoring residue F167
to insert deeper into the bilayer than the corresponding methionine
in Syt-1 (M173). By performing additional replica exchange umbrella
sampling calculations, we demonstrate that these additional contacts
increase the energetic cost of unbinding the Syt-7 C2 domains from
the bilayer, causing them to unbind more slowly than their counterparts
in Syt-1