This study explores the structural and electronic factors affecting the absorption spectra of 5-carboxy-tetramethylrhodamine (TAMRA) in water, a widely used fluorophore in imaging and molecular labeling in biophysical studies. Through molecular dynamics (MD) simulations and density functional theory (DFT) calculations, we examine TAMRA UV absorption spectra together with TAMRA-labeled peptides (Arg9, Arg4, Lys9). We found that DFT calculations with different functionals underestimate TAMRA maximum UV absorption peak by ~100 nm, resulting in the maximum at ca. 450 nm instead of the experimental value of ca. 550 nm. However, incorporating MD simulation snapshots of TAMRA in water, the UV maximum peak shifts and is in close agreement with the experimental results due to the rotation of TAMRA N(CH3)2 groups, effectively captured in MD simulations. The method is used to estimate the UV absorption spectra of TAMRA-labeled peptides, matching experimental values.
- Klíčová slova
- UV absorption spectra, fluorescent probes, molecular dynamics simulations, time‐dependent density functional theory,
- MeSH
- fluorescenční barviva chemie MeSH
- peptidy * chemie MeSH
- rhodaminy * chemie MeSH
- simulace molekulární dynamiky * MeSH
- spektrofotometrie ultrafialová MeSH
- teorie funkcionálu hustoty * MeSH
- voda chemie MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- 5-carboxytetramethylrhodamine succinimidyl ester MeSH Prohlížeč
- fluorescenční barviva MeSH
- peptidy * MeSH
- rhodaminy * MeSH
- voda MeSH
A novel pheophorbide derivative, trimethyl-152-[L-aspartyl]pheophorbide a was synthesised and investigated for anti-tumor activity. The prepared photosensitizer had good absorption in the phototherapeutic window and high ROS yields. It exhibited excellent phototoxicity higher than reference compound m-THPC when irradiated by 650 nm light in vitro, and obvious photodynamic anti-tumor effect in vivo. It causes cellular apoptosis or necrosis under laser irradiation and localizes in mitochondria, lysosome, and endoplasmic reticulum. The observed high efficacy was rationalized by efficient introduction into the blood by facile transfer through membranes, which is supported by molecular dynamics simulation studies. This work provides a new candidate photosensitizer for anti-cancer treatment.
- Klíčová slova
- cancer, keyword 5, molecular dynamics, pheophorbide, photodynamic therapy,
- Publikační typ
- časopisecké články MeSH
The mitochondrial ADP/ATP carrier (AAC, ANT), a member of the SLC25 family of solute carriers, plays a critical role in transporting purine nucleotides (ATP and ADP) as well as protons across the inner mitochondrial membrane. However, the precise mechanism and physiological significance of proton transport by ADP/ATP carrier remain unclear. Notably, the presence of uncouplers-such as long-chain fatty acids (FA) or artificial compounds like dinitrophenol (DNP)-is essential for this process. We explore two potential mechanisms that describe ADP/ATP carrier as either (i) a proton carrier that functions in the presence of FA or DNP, or (ii) an anion transporter (FA- or DNP). In the latter case, the proton is translocated by the neutral form of FA, which carries it from the matrix to the intermembrane space (FA-cycling hypothesis). Our recent results support this hypothesis. We describe a four-step mechanism for the "sliding" of the FA anion from the matrix to the mitochondrial intermembrane space and discuss a possible generalization of this mechanism to other SLC25 carriers.
- Klíčová slova
- MD simulations, bilayer lipid membranes, membrane proteins, mitochondrial transporter, reconstituted protein, uncoupling protein,
- MeSH
- 2,4-dinitrofenol farmakologie metabolismus MeSH
- iontový transport MeSH
- lidé MeSH
- mastné kyseliny metabolismus MeSH
- mitochondriální ADP/ATP-translokasy * metabolismus chemie MeSH
- mitochondriální membrány metabolismus MeSH
- mitochondrie metabolismus MeSH
- protony * MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- přehledy MeSH
- Názvy látek
- 2,4-dinitrofenol MeSH
- mastné kyseliny MeSH
- mitochondriální ADP/ATP-translokasy * MeSH
- protony * MeSH
The atmospheric partitioning of trifluoroacetic acid (TFA) in aerosol is a complex function of the size of suspended water droplets and their pH value. The unraveling of the affinity of TFA towards basic but not acidic conditions may be accomplished by providing an insight into the hydration pattern of undissociated TFA. Owing to rather scarce details on very dilute aqueous solutions of trifluoroacetic acid (TFA), we examined CF3COOD and CF3COONa solutions in D2O in the concentration range 0.001-0.1 mol dm-3 using transmission FTIR spectroscopy and computational methods. Besides detecting the signals originated from undissociated species in both CF3COOD (1787 cm-1 and 1766 cm-1 at c0 = 0.1 mol dm-3) and CF3COONa (1807 cm-1 at c0 = 0.1 mol dm-3) D2O solutions, through computational techniques we identified different TFA hydrates that contribute to the complexity of the spectral appearance. The combination of experimental and computational data suggested the concentration dependence of the predominant hydrogen bonding pattern of TFA. The results obtained in this work should help in understanding the partitioning of TFA into micron-size water droplets in the atmosphere in molecular and structural terms, i.e. the eventual stability of a hydrated complex for a particular TFA conformer.
The transformation of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) lipid bilayers from the gel (Lβ') to the fluid (Lα) phase involves an intermediate ripple (Pβ') phase forming a few degrees below the main transition temperature (Tm). While the exact cause of bilayer rippling is still debated, the presence of amphiphilic molecules, pH, and lipid bilayer architecture are all known to influence (pre)transition behavior. In particular, fatty acid chains interact with hydrophobic lipid tails, while the carboxylic groups simultaneously participate in proton transfer with interfacial water in the polar lipid region which is controlled by the pH of the surrounding aqueous medium. The molecular-level variations in the DPPC ripple phase in the presence of 2% palmitic acid (PA) were studied at pH levels 4.0, 7.3, and 9.1, where PA is fully protonated, partially protonated, or fully deprotonated. Bilayer thermotropic behavior was investigated by differential scanning calorimetry (DSC) and Fourier-transform infrared (FTIR) spectroscopy which agreed in their characterization of (pre)transition at pH of 9.1, but not at pH 4.0 and especially not at 7.3. Owing to the different insertion depths of protonated and deprotonated PA, along with the ability of protonated PA to undergo flip-flop in the bilayer, these two forms of PA show a different hydration pattern in the interfacial water layer. Finally, these results demonstrated the hitherto undiscovered potential of FTIR spectroscopy in the detection of the events occurring at the surface of lipid bilayers that obscure the low-cooperativity phase transition explored in this work.
- Klíčová slova
- 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) lipid bilayers, DSC and FTIR spectroscopy, Palmitic acid (PA), Ripple phase, pH-dependent (de)protonation,
- MeSH
- 1,2-dipalmitoylfosfatidylcholin * analogy a deriváty MeSH
- diferenciální skenovací kalorimetrie MeSH
- koncentrace vodíkových iontů MeSH
- kyselina palmitová * chemie MeSH
- lipidové dvojvrstvy * chemie MeSH
- molekulární struktura MeSH
- protony MeSH
- spektroskopie infračervená s Fourierovou transformací MeSH
- teplota MeSH
- změna skupenství MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- 1,2-dipalmitoylfosfatidylcholin * MeSH
- colfosceril palmitate MeSH Prohlížeč
- kyselina palmitová * MeSH
- lipidové dvojvrstvy * MeSH
- protony MeSH
This study employs molecular dynamics (MD) simulations to investigate the adsorption and aggregation behavior of simple polyarginine cell-penetrating peptides (CPPs), specifically modeled as R9 peptides, at zwitterionic phosphocholine POPC membranes under varying ionic strengths of two peptide concentrations and two concentrations of NaCl and CaCl2. The results reveal an intriguing phenomenon of R9 aggregation at the membrane, which is dependent on the ionic strength, indicating a salting-out effect. As the peptide concentration and ionic strength increase, peptide aggregation also increases, with aggregate lifetimes and sizes showing a corresponding rise, accompanied by the total decrease of adsorbed peptides at the membrane surface. Notably, in high ionic strength environments, large R9 aggregates, such as octamers, are also observed occasionally. The salting-out, typically uncommon for short positively charged peptides, is attributed to the unique properties of arginine amino acid, specifically by its side chain containing amphiphilic guanidinium (Gdm+) ion which makes both intermolecular hydrophobic like-charge Gdm+ - Gdm+ and salt-bridge Gdm+ - C-terminus interactions, where the former are increased with the ionic strength, and the latter decreased due to electrostatic screening. The aggregation behavior of R9 peptides at membranes can also be linked to their CPP translocation properties, suggesting that aggregation may aid in translocation across cellular membranes.
- Klíčová slova
- Ionic strength, Molecular dynamics simulations, Peptide aggregation, Phosphocholine lipid bilayers, Polyarginines, Salting-out,
- Publikační typ
- časopisecké články MeSH
Neutron scattering and molecular dynamics studies were performed on a concentrated aqueous tetramethylammonium (TMA) chloride solution to gain insight into the hydration shell structure of TMA, which is relevant for understanding its behavior in biological contexts of, e.g., properties of phospholipid membrane headgroups or interactions between DNA and histones. Specifically, neutron diffraction with isotopic substitution experiments were performed on TMA and water hydrogens to extract the specific correlation between hydrogens in TMA (HTMA) and hydrogens in water (HW). Classical molecular dynamics simulations were performed to help interpret the experimental neutron scattering data. Comparison of the hydration structure and simulated neutron signals obtained with various force field flavors (e.g. overall charge, charge distribution, polarity of the CH bonds and geometry) allowed us to gain insight into how sensitive the TMA hydration structure is to such changes and how much the neutron signal can capture them. We show that certain aspects of the hydration, such as the correlation of the hydrogen on TMA to hydrogen on water, showed little dependence on the force field. In contrast, other correlations, such as the ion-ion interactions, showed more marked changes. Strikingly, the neutron scattering signal cannot discriminate between different hydration patterns. Finally, ab initio molecular dynamics was used to examine the three-dimensional hydration structure and thus to benchmark force field simulations. Overall, while neutron scattering has been previously successfully used to improve force fields, in the particular case of TMA we show that it has only limited value to fully determine the hydration structure, with other techniques such as ab initio MD being of a significant help.
- Publikační typ
- časopisecké články MeSH
Aqueous solutions of acetic acid (AA) have been intensively explored for decades with a particular attention addressed to the hydrogen bond network generated by COOH group at different concentrations. In majority of studies conducted so far the envelope originated from νCO is decomposed into two bands assigned to differently hydrated monomers: the one presumably to AA···H2O, and another one to AA···(H2O)2. In order to examine if species other than the mentioned monomers produce this spectral signature, we performed computational and FTIR spectroscopic study of AA in aqueous solutions. Dilute solutions of deuterated acetic acid (CD3COOD) in D2O and in C2Cl4 as a reference were prepared (c0 = 0.001, 0.01 and 0.1 mol dm-3) as well as of deuterated sodium acetate (CD3COONa) in D2O. CD3COOD in 0.1 mol dm-3 solution in D2O displays a feature that separated in two signals with maxima at 1706 cm-1 and 1687 cm-1. A combined DFT and molecular dynamics study performed in this work showed the assignation of those spectral bands to be a more complex problem than previously thought, with syn-anti isomerism and hydration contributing to the experimentally observed broad νCO envelope.
- Klíčová slova
- Acetic acid, Density functional theory, Dimerization, Dissociation, FTIR spectroscopy, Hydration, Molecular dynamics simulations, Sodium acetate, syn-anti isomerism,
- Publikační typ
- časopisecké články MeSH
Adsorption of cell-penetrating peptides (CPPs) at cellular membranes is the first and necessary step for their subsequent translocation across cellular membranes into the cytosol. It has been experimentally shown that CPPs rich in arginine (Arg) amino acid penetrate across phospholipid bilayers more effectively than their lysine (Lys) rich counterparts. In this work, we aim to understand the differences in the first translocation step, adsorption of Arg9 and Lys9 peptides at fully hydrated neutral phosphatidylcholine (PC) and phosphatidylethanolamine (PE) lipid bilayers and evaluate in detail the energetics of the process using molecular dynamics (MD) simulations and free energy calculations of adsorption of the single peptide. We show that the adsorption of Arg9 is energetically feasible, with the free energy of adsorption being ∼-5.0 kcal mol-1 at PC and ∼-5.5 kcal mol-1 at PE bilayers. In contrast, adsorption of Lys9 is not observed at PC bilayers, and their adsorption at PE bilayers is very weak, being ∼-0.5 kcal mol-1. We show by energy decomposition and analysis of peptide hydration along the membrane that significantly stronger electrostatic interactions of Arg9 with lipid phosphate groups, together with the greater loss of peptide hydration (and in turn stronger hydrophobic interactions) along the membrane translocation path, are the main driving factors governing the adsorption of Arg-rich peptides at neutral lipid bilayers in contrast to Lys-rich peptides. Finally, we also compare the energetics in lipid/bilayer systems with the density functional theory (DFT) calculations of the corresponding model systems in the continuum water model and reveal the energetic differences in different environments.
- MeSH
- fosfatidylcholiny chemie MeSH
- fosfolipidy MeSH
- lipidové dvojvrstvy chemie MeSH
- penetrační peptidy * chemie MeSH
- polylysin * MeSH
- simulace molekulární dynamiky MeSH
- teorie funkcionálu hustoty MeSH
- termodynamika MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- fosfatidylcholiny MeSH
- fosfolipidy MeSH
- lipidové dvojvrstvy MeSH
- penetrační peptidy * MeSH
- polyarginine MeSH Prohlížeč
- polylysin * MeSH
Mitochondrial adenine nucleotide translocase (ANT) exchanges ADP for ATP to maintain energy production in the cell. Its protonophoric function in the presence of long-chain fatty acids (FA) is also recognized. Our previous results imply that proton/FA transport can be best described with the FA cycling model, in which protonated FA transports the proton to the mitochondrial matrix. The mechanism by which ANT1 transports FA anions back to the intermembrane space remains unclear. Using a combined approach involving measurements of the current through the planar lipid bilayers reconstituted with ANT1, site-directed mutagenesis and molecular dynamics simulations, we show that the FA anion is first attracted by positively charged arginines or lysines on the matrix side of ANT1 before moving along the positively charged protein-lipid interface and binding to R79, where it is protonated. We show that R79 is also critical for the competitive binding of ANT1 substrates (ADP and ATP) and inhibitors (carboxyatractyloside and bongkrekic acid). The binding sites are well conserved in mitochondrial SLC25 members, suggesting a general mechanism for transporting FA anions across the inner mitochondrial membrane.
- Klíčová slova
- AAC, ADP/ATP carrier, arachidonic acid, fatty acid cycling hypothesis, fatty acids anion transport, proton transport, uncoupling proteins,
- MeSH
- adenosintrifosfát metabolismus MeSH
- anionty metabolismus MeSH
- lipidové dvojvrstvy * MeSH
- mastné kyseliny metabolismus MeSH
- mitochondriální ADP/ATP-translokasy metabolismus MeSH
- protony * MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- adenosintrifosfát MeSH
- anionty MeSH
- lipidové dvojvrstvy * MeSH
- mastné kyseliny MeSH
- mitochondriální ADP/ATP-translokasy MeSH
- protony * MeSH
