Molecular Modeling of Biological Systems: from a Force Field Study to Modeling of an Anti-Microbial Peptide in Water and a Complex S. aureus Membrane

Interest in studying anti-microbial peptides (AMPs) has increased because of their potential as a future applicable antibiotic drug. In my research under supervision of Professor Greenfield, we attempt to understand the interaction of AMPs against membrane of bacteria. We are studying the structure of two novel hybrid AMPs LM7-1 and LM7-2 that were designed previously in Professor Martin’s research group (Cell and Molecular Biology Department). These AMPs differ in sequence only at the 15th residue. Several experimental studies have been successfully investigated mechanisms of AMPs against bacteria, but there is still much uncertainty in the exact mechanisms because of experimental restrictions. Since molecular modeling provides atomistic details and 3D structure of a system, it can significantly contribute to investigating these mechanisms in more detail. In this talk, I will show how molecular modeling helps in understanding complex systems and deciphering biochemistry information that it is impossible to obtain by experiments. And since the precision of biomolecular modeling results is significantly based on force field (FF) parameters, the importance of developing FFs will be demonstrated in this talk. Force Field development. Our simulation results on the basis of CHARMM36 (C36) FF show that the structures of two aromatic amino acids, Tryptophan (Trp) and Tyrosine (Tyr), deviate from planarity. Hence, we investigated the geometry, dynamics, and out-of-plane vibrations of atoms in these rings by imposing improper torsion and changing torsion angle force constants. To that end, molecular dynamics (MD) simulation and all-atom normal mode analysis (NMA) were implemented. We could match the pattern and frequencies of out-of-plane vibrations of these rings with Raman and infrared spectra, and decrease the extent of out-of-plane vibrations for atoms in these rings. A Helical Peptide in water. AMPs usually have a helical structure on the membrane of bacteria. Some studies have stated that the flexible loop at the middle of helical AMPs, which leads peptide to bend and snap the lipid bilayer, has a direct effect on AMPs activity against bacteria. We computationally studied dynamics and vibrations of a helical and a helix-hinged-helix structure of a LM7-2 in solution. Although some vibrational experimental studies have been done on proteins or peptides, we show extended and interesting details about peptide vibrations by our study. We applied instantaneous NMA and Fourier Transform method to understand how a change in the structure of a peptide will affect peptide fluctuations. Raman and infrared spectra cannot indicate these motions that correspond to the low intensity measured frequencies. A complex lipid membrane. Lipid bilayers play a crucial role in a peptide-membrane interactions. Therefore, more real system of lipids will provide more detail of this interaction. To that end, we have designed the most realistic S. aureus membrane by including 19 different types of lipids, compared to other simulations that implemented a range of 2-5 different lipids. We applied Reverse Monte Carlo method to match lipid bilayer composition to experimental results in the literature. Dynamics and membrane characteristics of this complex system were studied

Planarity and out-of-plane vibrational modes of tryptophan and tyrosine in biomolecular modeling

Tryptophan and tyrosine are aromatic amino acids that play significant roles in the folding processes of proteins at water-membrane interfaces because of their amphipathic structures. Employing appropriate heteroaromatic molecular structures are essential for obtaining accurate dynamics and predictive capabilities in molecular simulations of these amino acids. In this study, molecular dynamics simulations that applied the most recent version of the CHARMM36 force field were conducted on aqueous solutions of tryptophan and of tyrosine. Geometric analysis and dynamics quantified how aromatic rings deviated from planar structures and exhibited out-of-plane fluctuations. Radial distribution functions showed possible biological significance because extent of ring planarity slightly affected local water concentrations near aromatic rings. Instantaneous all-atom normal mode analysis (NMA) and Fourier transformation of time autocorrelation functions of out-of-plane displacements were applied to study out-of-plane vibrations of atoms in those rings. NMA started with minimum energy configurations and then averaged over fluctuations in aqueous solution. The frequencies and frequency patterns that were obtained for tryptophan and tyrosine with CHARMM36 differed from literature reports of Raman spectra, infrared spectra, and frequencies calculated using quantum mechanics, with some out-of-plane modes found at higher frequencies. Effects of imposing improper torsion potentials and changing torsion angle force constants were investigated for all atoms in the rings of tryptophan and tyrosine. Results show that these coarse force fields variations only affect planarity and out-of-plane vibrations of atoms within the rings, not other vibrations. Although increasing improper torsion force constants reduced deviations from aromatic ring planarity significantly, it increased out-of-plane mode frequencies. Reducing torsion angle force constants (with and without improper torsions) shifted modes to lower frequencies. A combination of decreasing most torsion angle force constants for ring atoms in both amino acids and including improper torsion forces attained frequencies and frequency patterns for out-of-plane normal modes that were more similar to literature spectra. These force field variations decreased the extents of out-of-plane vibrations within the heteroaromatic rings of tryptophan, especially around the nitrogen atom in the ring, but not within the heteroaromatic ring of tyrosine. Conclusions were unaffected by peptide endgroup, water, or simulation ensemble

Computing Individual Area per Head Group Reveals Lipid Bilayer Dynamics

Lipid bilayers express a range of phases from solid-like to gel-like to liquid-like as a function of temperature and lipid surface concentration. The area occupied per lipid head group serves as one useful indicator of the bilayer phase, in conjunction with the two-dimensional radial distribution function (i.e., structure factor) within the bilayer. Typically, the area per head group is determined by dividing the bilayer area equally among all head groups. Such an approach is less satisfactory for a multicomponent set of diverse lipids. In this work, area determination is performed on a lipid-by-lipid basis by attributing to a lipid the volume that surrounds each atom. Voronoi tessellation provides this division of the interfacial region on a per-atom basis. The method is applied to a multicomponent system of water, NaCl, and 19 phospholipid types that was devised recently [Langmuir 2022, 38, 9481–9499] as a computational representation of the Gram-positive Staphylococcus aureus phospholipid bilayer. Results demonstrate that lipids and water molecules occupy similar extents of area within the interfacial region; ascribing area only to head groups implicitly incorporates assumptions about head group hydration. Results further show that lipid tails provide non-negligible contributions to area on the membrane side of the bilayer–water interface. Results for minimum and maximum area of individual lipids reveal that spontaneous fluctuations displace head groups more than 10 Å from the interfacial region during an NPT simulation at 310 K, leading to a zero contribution to total area at some times. Total area fluctuations and fluctuations per individual lipid relax with a correlation time of ∼10 ns. The method complements density profile as an approach to quantify the structure and dynamics of computational lipid bilayers

Generation and Computational Characterization of a Complex Staphylococcus aureus Lipid Bilayer

Studies indicate a crucial cell membrane role in the antibiotic resistance of Staphylococcus aureus. To simulate its membrane structure and dynamics, a complex molecular-scale computational representation of the S. aureus lipid bilayer was developed. Phospholipid types and their amounts were optimized by reverse Monte Carlo to represent characterization data from the literature, leading to 19 different phospholipid types that combine three headgroups [phosphatidylglycerol, lysyl-phosphatidylglycerol (LPG), and cardiolipin] and 10 tails, including iso- and anteiso-branched saturated chains. The averaged lipid bilayer thickness was 36.7 Å, and area per headgroup was 67.8 Å2. Phosphorus and nitrogen density profiles showed that LPG headgroups tended to be bent and oriented more parallel to the bilayer plane. The water density profile showed that small amounts reached the membrane center. Carbon density profiles indicated hydrophobic interactions for all lipids in the middle of the bilayer. Bond vector order parameters along each tail demonstrated different C–H ordering even within distinct lipids of the same type; however, all tails followed similar trends in average order parameter. These complex simulations further revealed bilayer insights beyond those attainable with monodisperse, unbranched lipids. Longer tails often extended into the opposite leaflet. Carbon at and beyond a branch showed significantly decreased ordering compared to carbon in unbranched tails; this feature arose in every branched lipid. Diverse tail lengths distributed these disordered methyl groups throughout the middle third of the bilayer. Distributions in mobility and ordering reveal diverse properties that cannot be obtained with monodisperse lipids

Generation and Computational Characterization of a Complex Staphylococcus aureus Lipid Bilayer

Studies indicate a crucial cell membrane role in the antibiotic resistance of Staphylococcus aureus. To simulate its membrane structure and dynamics, a complex molecular-scale computational representation of the S. aureus lipid bilayer was developed. Phospholipid types and their amounts were optimized by reverse Monte Carlo to represent characterization data from the literature, leading to 19 different phospholipid types that combine three headgroups [phosphatidylglycerol, lysyl-phosphatidylglycerol (LPG), and cardiolipin] and 10 tails, including iso- and anteiso-branched saturated chains. The averaged lipid bilayer thickness was 36.7 Å, and area per headgroup was 67.8 Å2. Phosphorus and nitrogen density profiles showed that LPG headgroups tended to be bent and oriented more parallel to the bilayer plane. The water density profile showed that small amounts reached the membrane center. Carbon density profiles indicated hydrophobic interactions for all lipids in the middle of the bilayer. Bond vector order parameters along each tail demonstrated different C–H ordering even within distinct lipids of the same type; however, all tails followed similar trends in average order parameter. These complex simulations further revealed bilayer insights beyond those attainable with monodisperse, unbranched lipids. Longer tails often extended into the opposite leaflet. Carbon at and beyond a branch showed significantly decreased ordering compared to carbon in unbranched tails; this feature arose in every branched lipid. Diverse tail lengths distributed these disordered methyl groups throughout the middle third of the bilayer. Distributions in mobility and ordering reveal diverse properties that cannot be obtained with monodisperse lipids

Computational Study of Helical and Helix-Hinge-Helix Conformations of an Anti-Microbial Peptide in Solution by Molecular Dynamics and Vibrational Analysis

Many classical antimicrobial peptides adopt an amphipathic helical structure at a water-membrane interface. Prior studies led to the hypothesis that a hinge near the middle of a helical peptide plays an important role in facilitating peptide-membrane interactions.Here, dynamics and vibrations of a designed hybrid antimicrobial peptide LM7-2 in solution were simulated to investigate its hinge formation.Molecular dynamics simulation results on the basis of the CHARMM36 force field showed that the $\alpha$-helix LM7-2 bent around two or three residues near the middle of the peptide, stayed in a helix-hinge-helix conformation for a short period of time, and then returned to a helical conformation.High resolution computational vibrational techniques were applied on the LM7-2 system when it has $\alpha$-helical and helix-hinge-helix conformations to understand how this structural change affects its inherent vibrations.These studies concentrated on the calculation of frequencies that correspond to backbone amide bands I, II, and III: vibrational modes that are sensitive to changes in the secondary structure of peptides and proteins.To that end, Fourier transforms were applied to thermal fluctuations in C-N-H angles, C-N bond lengths, and C=O bond lengths of each amide group. In addition, instantaneous all-atom normal mode analysis was applied to monitor and detect the characteristic amide bands of each amide group within LM7-2 during the MD simulation.Computational vibrational results indicate that shapes and frequencies of amide bands II and especially III were altered only for amide groups near the hinge.These methods provide high resolution vibrational information that can complement spectroscopic vibrational studies. They assist in interpreting spectra of similar systems and suggest a marker for the presence of the helix-hinge-helix motif. Moreover, radial distribution functions indicated an increase in the probability of hydrogen bonding between water and a hydrogen atom connected to nitrogen (HN) in such a hinge.The probability of intramolecular hydrogen bond formation between HN and an amide group oxygen atom within LM7-2 was lower around the hinge.No correlation has been found between the presence of a hinge andhydrogen bonds between amide group oxygen atoms and the hydrogen atoms of water molecules.This result suggests a mechanism for hinge formation wherein hydrogen bonds to oxygen atoms of water replace intramolecular hydrogen bonds as the peptide backbone folds