A mixed quantum–classical method is proposed to describe the dynamical behavior of a diatomic mol... more A mixed quantum–classical method is proposed to describe the dynamical behavior of a diatomic molecule in a gas environment. The vibrational coordinate is treated quantum-mechanically and all other degrees of freedom classically. Within the present approach the classical equations for the rotational motion have no singularities. A symplectic, energy conserving and time-reversible algorithm is used for the propagation. As an application we treat the dynamics of I2 molecules excited by femtosecond laser pulses moving in collision-free and high pressure rare-gas environments. For freely rotating I2 molecules, the thermal average over rotational states leads to the decay of the pump–probe signal. For I2 in inert gases, we show that dephasing by collisions with the buffer gas is a weak effect in comparison with the decay of the signal due to the anharmonicity of the potential energy curve. Therefore the oscillating structure of the pump–probe signals depends weakly on the mass and the pressure of the solvent, ...
Anoplin is a linear 10-amino acid amphipathic peptide (Gly-Leu-Leu-Lys-Arg-Ile-Lys-Thr-Leu-Leu-NH... more Anoplin is a linear 10-amino acid amphipathic peptide (Gly-Leu-Leu-Lys-Arg-Ile-Lys-Thr-Leu-Leu-NH2) derived from the venom sac of the solitary wasp. It has broad antimicrobial activity, including an antibacterial one. However, the inhibition of bacterial growth requires several dozen micromolar concentrations of this peptide. Anoplin is positively charged and directly interacts with anionic biological membranes forming an α-helix that disrupts the lipid bilayer. To improve the bactericidal properties of anoplin by stabilizing its helical structure, we designed and synthesized its analogs with hydrocarbon staples. The staple was introduced at two locations resulting in different charges and amphipathicity of the analogs. Circular dichroism studies showed that all modified anoplins adopted an α-helical conformation, both in the buffer and in the presence of membrane mimics. As the helicity of the stapled anoplins increased, their stability in trypsin solution improved. Using the propi...
An entry from the Cambridge Structural Database, the world's repository for small molecule cr... more An entry from the Cambridge Structural Database, the world's repository for small molecule crystal structures. The entry contains experimental data from a crystal diffraction study. The deposited dataset for this entry is freely available from the CCDC and typically includes 3D coordinates, cell parameters, space group, experimental conditions and quality measures.
Ó The Author(s) 2010. This article is published with open access at Springerlink.com Abstract Tra... more Ó The Author(s) 2010. This article is published with open access at Springerlink.com Abstract Translation on the ribosome is controlled by external factors. During polypeptide lengthening, elongation factors EF-Tu and EF-G consecutively interact with the bacterial ribosome. EF-Tu binds and delivers an aminoacyl-tRNA to the ribosomal A site and EF-G helps translocate the tRNAs between their binding sites after the peptide bond is formed. These processes occur at the expense of GTP. EF-Tu:tRNA and EF-G are of similar shape, share a common binding site, and undergo large conformational changes on interaction with the ribosome. To characterize the internal motion of these two elongation factors, we used 25 ns long all-atom molecular dynamics simulations. We observed enhanced mobility of EF-G domains III, IV, and V and of tRNA in the EF-Tu:tRNA complex. EF-Tu:GDP complex acquired a configuration different from that found in the crystal structure of EF-Tu with a GTP analogue, showing conf...
Lincosamides are a class of antibiotics used both in clinical and veterinary practice for a wide ... more Lincosamides are a class of antibiotics used both in clinical and veterinary practice for a wide range of pathogens. This group of drugs inhibits the activity of the bacterial ribosome by binding to the 23S RNA of the large ribosomal subunit and blocking protein synthesis. Current-ly, three X-ray structures of the ribosome in complex with clindamycin are available in the Protein Data Bank, which reveal that there are two distinct conformations of the pyrrolidinyl propyl group of the bound clindamycin. In this work, we used quantum mechanical methods to investigate the probable conformations of clindamycin in order to explain the two binding modes in the ribosomal 23S RNA. We studied three lincosamide antibiotics: clindamycin, lincomycin, and pirlimycin at the B3LYP level with the 6-31G* * basis set. The focus of our work was to connect the conformational landscape and electron densities of the two clindamycin conformers found experimentally with their physicochemical properties. For...
Using molecular dynamics simulations, we describe how crowded environments affect the internal dy... more Using molecular dynamics simulations, we describe how crowded environments affect the internal dynamics and diffusion of the hepatitis C virus proteases NS3/4A. This protease plays a key role in viral replication and is successfully used as a target for antiviral treatment. The NS3 enzyme requires a peptide cofactor, called NS4A, with its central part interacting with the NS3 b-sheet, and flexible, protruding terminal tails that are unstructured in water solution. The simulations describe the enzyme and water molecules at atomistic resolution, whereas crowders are modeled via either all-atom or coarse-grained models to emphasize different aspects of crowding. Crowders reflect the polyethylene glycol (PEG) molecules used in the experiments to mimic the crowded surrounding. A bead-shell model of folded coarse-grained PEG molecules considers mainly the excluded volume effect, whereas all-atom PEG models afford more protein-like crowder interactions. Circular dichroism spectroscopy experiments of the NS4A N-terminal tail show that a helical structure is formed in the presence of PEG crowders. The simulations suggest that crowding may assist in the formation of an NS4A helical fragment, positioned exactly where a transmembrane helix would fold upon the NS4A contact with the membrane. In addition, partially interactive PEGs help the NS4A N-tail to detach from the protease surface, thus enabling the process of helix insertion and potentially helping the virus establish a replication machinery needed to produce new viruses. Results point to an active role of crowding in assisting structural changes in disordered protein fragments that are necessary for their biological function.
The identification of novel targets for antimicrobial agents is crucial for combating infectious ... more The identification of novel targets for antimicrobial agents is crucial for combating infectious diseases caused by evolving bacterial pathogens. Components of bacterial toxin–antitoxin (TA) systems have been recognized as promising therapeutic targets. These widespread genetic modules are usually composed of two genes that encode a toxic protein targeting an essential cellular process and an antitoxin that counteracts the activity of the toxin. Uncontrolled toxin expression may elicit a bactericidal effect, so they may be considered “intracellular molecular bombs” that can lead to elimination of their host cells. Based on the molecular nature of antitoxins and their mode of interaction with toxins, TA systems have been classified into six groups. The most prevalent are type II TA systems. Due to their ubiquity among clinical isolates of pathogenic bacteria and the essential processes targeted, they are promising candidates for the development of novel antimicrobial strategies. In t...
Short oligonucleotides binding complementary mRNA essential for bacterial growth could be used as... more Short oligonucleotides binding complementary mRNA essential for bacterial growth could be used as antibacterials provided that they are delivered to bacterial cells. One promising oligonucleotide is peptide nucleic acid (PNA), a synthetic DNA analog with a neutral peptide-like backbone. Unfortunately, bacteria do not import short oligonucleotides, including PNA, from the environment and efficient non-invasive methods for PNA delivery have not been found. To deliver PNA to bacterial cells, we have used vitamin B 12. Most bacteria need to import vitamin B 12 from the environment to sustain life. We have shown that vitamin B 12 covalently linked with PNA, delivers PNA to E. coli cells. Free vitamin B 12 enters E. coli via the TonB-dependent transport system and is recognized by the outer-membrane vitamin B 12-specific BtuB receptor. We engineered the E. coli DbtuB mutant and found that transport of the vitamin B 12-PNA conjugate also requires BtuB. Thus, vitamin B 12 uses the BtuB receptor to deliver PNA through the E. coli outer membrane. Overall, we have proven that vitamin B 12 transport system can be hijacked for PNA delivery to E. coli cells. Further, using Gaussian-force simulated annealing molecular dynamics with umbrella sampling we determined the mechanism of vitamin B 12-PNA permeation through the BtuB receptor protein. BtuB is a beta-barrel occluded by its luminal domain, but we found that partial unfolding of this domain makes the passage of the conjugate mechanically feasible. PNA movement into the beta-barrel is energetically favorable since, while inside BtuB, PNA extends. In addition, we found that BtuB extracellular loops are actively involved in transport through an induced-fit mechanism.
Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature, 2017
Binding of metal ions is an important factor governing the folding and dynamics of RNA. Shielding... more Binding of metal ions is an important factor governing the folding and dynamics of RNA. Shielding of charges in the polyanionic backbone allows RNA to adopt a diverse range of folded structures that give rise to their many functions within the cell. Some RNA sequences fold only in the presence of Mg 2+ , which may be bound via direct interactions or occupy the more diffuse "ion atmosphere" around the RNA. To understand the driving forces for RNA folding, it is important to be able to fully characterize the distribution of metal ions around the RNA. In this work, a combined Grand Canonical Monte Carlo-Molecular Dynamics (GCMC-MD) method is applied to characterize Mg 2+ distributions around folded RNA structures. The GCMC-MD approach identifies known innerand outer-shell Mg 2+ coordination, while also predicting new regions occupied by Mg 2+ that are not observed in crystal structures but that may be relevant in solution, including the case of the Mg 2+ riboswitch, for which alternate Mg 2+ binding sites may have implications for its function. This work represents a significant step forward in establishing a structural and thermodynamic description of RNA−Mg 2+ interactions and their role in RNA structure and function.
Peptide nucleic acids are promising nucleic acid analogs for antisense therapies as they can form... more Peptide nucleic acids are promising nucleic acid analogs for antisense therapies as they can form stable duplex and triplex structures with DNA and RNA. Computational studies of PNA-containing duplexes and triplexes are an important component for guiding their design, yet existing force fields have not been well validated and parameterized with modern computational capabilities. We present updated CHARMM and Amber force fields for PNA that greatly improve the stability of simulated PNAcontaining duplexes and triplexes in comparison with experimental structures and allow such systems to be studied on microsecond time scales. The force field modifications focus on reparametrized PNA backbone torsion angles to match high-level quantum mechanics reference energies for a model compound. The microsecond simulations of PNA-PNA, PNA-DNA, PNA-RNA, and PNA-DNA-PNA complexes also allowed a comprehensive analysis of hydration and ion interactions with such systems.
The widespread emergence of bacterial resistance to existing antibiotics forces the development o... more The widespread emergence of bacterial resistance to existing antibiotics forces the development of new therapeutic agents. The use of short modified oligonucleotides, such as peptide nucleic acids (PNAs), seems a promising strategy. However, the uptake of such oligonucleotides is limited by the bacterial cell wall and is species-dependent. Therefore, new carriers for PNAs should be extensively explored. In this study, we examined the antibacterial activity of vitamin B 12 −PNA conjugates. Vitamin B 12 was covalently linked to a PNA oligomer targeted at the mRNA of an essential acpP gene encoding acyl carrier protein in Escherichia coli. PNA−vitamin B 12 conjugates were synthesized using the Cu(I)-catalyzed 1,3-dipolar cycloaddition. We examined two types of linkers between vitamin B 12 and PNA, including a cleavable disulfide bond. As a positive control for PNA uptake, we used PNA conjugated to the most widely used cell-penetrating peptide (KFF) 3 K. We found that vitamin B 12 −PNA conjugates inhibit E. coli growth at a concentration of 5 μM, similar as (KFF) 3 K−PNA. We also showed that vitamin B 12 −PNA conjugates are stable in the presence of biological media. This study provides the foundation for improving and developing PNAs conjugated to vitamin B 12 as antibacterials.
Clindamycin, a lincosamide antibiotic, binds to 23S ribosomal RNA and inhibits protein synthesis.... more Clindamycin, a lincosamide antibiotic, binds to 23S ribosomal RNA and inhibits protein synthesis. The A2058G mutation in 23S RNA results in bacterial resistance to clindamycin. To understand the influence of this mutation on short-range interactions of clindamycin with 23S RNA, we carried out full-atom molecular dynamics simulations of a ribosome fragment containing clindamycin binding site. We compared the dynamical behavior of this fragment simulated with and without the A2058G mutation. Molecular dynamics simulations suggest that clindamycin in the native ribosomal binding site is more internally flexible than in the A2058G mutant. Only in the native ribosome fragment did we observe intramolecular conformational change of clindamycin around its C7-N1-C10-C11 dihedral. In the mutant, G2058 makes more stable hydrogen bonds with clindamycin hindering its conformational freedom in the ribosome-bound state. Clindamycin binding site is located in the entrance to the tunnel through which the newly synthesized polypeptide leaves the ribosome. We observed that in the native ribosome fragment, clindamycin blocks the passage in the tunnel entrance, whereas in the mutated fragment the aperture is undisturbed due to a different mode of binding of clindamycin in the mutant. Restricted conformational freedom of clindamycin in a position not blocking the tunnel entrance in the A2058G mutant could explain the molecular mechanism of bacterial resistance against clindamycin occurring in this mutant.
Biochimica et Biophysica Acta (BBA) - Biomembranes, 2019
Proteinase 3 (PR3), together with other serine proteases, such as neutrophil elastase (NE) and ca... more Proteinase 3 (PR3), together with other serine proteases, such as neutrophil elastase (NE) and cathepsin G (CG), regulates inflammatory and immune responses. However, in comparison with NE and CG, there is increasing evidence that PR3 functions significantly differ. In particular, PR3 can bind to cell membranes and such membrane-bound PR3 (mbPR3) might be differently involved in the activation of cytokines, growth factors, cellular receptors, and in the regulation of cell apoptosis. For instance, PR3 membrane binding can block some "eat me" signals, notably, phosphatidylserine membrane lipid, and facilitate non-resolving inflammation. Based on the clear evidence that PR3 membrane binding affects the biological functions of PR3, we designed peptidomimetic inhibitors that can remove mbPR3 from the membrane surface in vitro without influencing PR3 catalytic activity. Such inhibitors, which specifically target PR3 binding to membranes, are still lacking. In particular, we found peptidomimetics that inhibit binding of PR3 to POPC:PS liposomes, which mimic the biological environment of PR3.
Journal of Chemical Information and Modeling, 2019
Transport of various molecules facilitated with membrane proteins is necessary for maintaining ho... more Transport of various molecules facilitated with membrane proteins is necessary for maintaining homeostasis in living cells. In humans, dysfunction of these proteins leads to many diseases. Thus, understanding how the membrane proteins function may help using them as therapeutic targets. To successfully investigate the mechanistic aspects of transport, the choice of appropriate methods is crucial. We review the computational methods that have proven most effective in investigating transport events, specifically, deterministic time-dependent classical molecular dynamics and its enhanced sampling variants, as well as methods based on Brownian dynamics. We describe technical aspects of these methods and examples of their novel variants or combinations that have
A mixed quantum–classical method is proposed to describe the dynamical behavior of a diatomic mol... more A mixed quantum–classical method is proposed to describe the dynamical behavior of a diatomic molecule in a gas environment. The vibrational coordinate is treated quantum-mechanically and all other degrees of freedom classically. Within the present approach the classical equations for the rotational motion have no singularities. A symplectic, energy conserving and time-reversible algorithm is used for the propagation. As an application we treat the dynamics of I2 molecules excited by femtosecond laser pulses moving in collision-free and high pressure rare-gas environments. For freely rotating I2 molecules, the thermal average over rotational states leads to the decay of the pump–probe signal. For I2 in inert gases, we show that dephasing by collisions with the buffer gas is a weak effect in comparison with the decay of the signal due to the anharmonicity of the potential energy curve. Therefore the oscillating structure of the pump–probe signals depends weakly on the mass and the pressure of the solvent, ...
Anoplin is a linear 10-amino acid amphipathic peptide (Gly-Leu-Leu-Lys-Arg-Ile-Lys-Thr-Leu-Leu-NH... more Anoplin is a linear 10-amino acid amphipathic peptide (Gly-Leu-Leu-Lys-Arg-Ile-Lys-Thr-Leu-Leu-NH2) derived from the venom sac of the solitary wasp. It has broad antimicrobial activity, including an antibacterial one. However, the inhibition of bacterial growth requires several dozen micromolar concentrations of this peptide. Anoplin is positively charged and directly interacts with anionic biological membranes forming an α-helix that disrupts the lipid bilayer. To improve the bactericidal properties of anoplin by stabilizing its helical structure, we designed and synthesized its analogs with hydrocarbon staples. The staple was introduced at two locations resulting in different charges and amphipathicity of the analogs. Circular dichroism studies showed that all modified anoplins adopted an α-helical conformation, both in the buffer and in the presence of membrane mimics. As the helicity of the stapled anoplins increased, their stability in trypsin solution improved. Using the propi...
An entry from the Cambridge Structural Database, the world's repository for small molecule cr... more An entry from the Cambridge Structural Database, the world's repository for small molecule crystal structures. The entry contains experimental data from a crystal diffraction study. The deposited dataset for this entry is freely available from the CCDC and typically includes 3D coordinates, cell parameters, space group, experimental conditions and quality measures.
Ó The Author(s) 2010. This article is published with open access at Springerlink.com Abstract Tra... more Ó The Author(s) 2010. This article is published with open access at Springerlink.com Abstract Translation on the ribosome is controlled by external factors. During polypeptide lengthening, elongation factors EF-Tu and EF-G consecutively interact with the bacterial ribosome. EF-Tu binds and delivers an aminoacyl-tRNA to the ribosomal A site and EF-G helps translocate the tRNAs between their binding sites after the peptide bond is formed. These processes occur at the expense of GTP. EF-Tu:tRNA and EF-G are of similar shape, share a common binding site, and undergo large conformational changes on interaction with the ribosome. To characterize the internal motion of these two elongation factors, we used 25 ns long all-atom molecular dynamics simulations. We observed enhanced mobility of EF-G domains III, IV, and V and of tRNA in the EF-Tu:tRNA complex. EF-Tu:GDP complex acquired a configuration different from that found in the crystal structure of EF-Tu with a GTP analogue, showing conf...
Lincosamides are a class of antibiotics used both in clinical and veterinary practice for a wide ... more Lincosamides are a class of antibiotics used both in clinical and veterinary practice for a wide range of pathogens. This group of drugs inhibits the activity of the bacterial ribosome by binding to the 23S RNA of the large ribosomal subunit and blocking protein synthesis. Current-ly, three X-ray structures of the ribosome in complex with clindamycin are available in the Protein Data Bank, which reveal that there are two distinct conformations of the pyrrolidinyl propyl group of the bound clindamycin. In this work, we used quantum mechanical methods to investigate the probable conformations of clindamycin in order to explain the two binding modes in the ribosomal 23S RNA. We studied three lincosamide antibiotics: clindamycin, lincomycin, and pirlimycin at the B3LYP level with the 6-31G* * basis set. The focus of our work was to connect the conformational landscape and electron densities of the two clindamycin conformers found experimentally with their physicochemical properties. For...
Using molecular dynamics simulations, we describe how crowded environments affect the internal dy... more Using molecular dynamics simulations, we describe how crowded environments affect the internal dynamics and diffusion of the hepatitis C virus proteases NS3/4A. This protease plays a key role in viral replication and is successfully used as a target for antiviral treatment. The NS3 enzyme requires a peptide cofactor, called NS4A, with its central part interacting with the NS3 b-sheet, and flexible, protruding terminal tails that are unstructured in water solution. The simulations describe the enzyme and water molecules at atomistic resolution, whereas crowders are modeled via either all-atom or coarse-grained models to emphasize different aspects of crowding. Crowders reflect the polyethylene glycol (PEG) molecules used in the experiments to mimic the crowded surrounding. A bead-shell model of folded coarse-grained PEG molecules considers mainly the excluded volume effect, whereas all-atom PEG models afford more protein-like crowder interactions. Circular dichroism spectroscopy experiments of the NS4A N-terminal tail show that a helical structure is formed in the presence of PEG crowders. The simulations suggest that crowding may assist in the formation of an NS4A helical fragment, positioned exactly where a transmembrane helix would fold upon the NS4A contact with the membrane. In addition, partially interactive PEGs help the NS4A N-tail to detach from the protease surface, thus enabling the process of helix insertion and potentially helping the virus establish a replication machinery needed to produce new viruses. Results point to an active role of crowding in assisting structural changes in disordered protein fragments that are necessary for their biological function.
The identification of novel targets for antimicrobial agents is crucial for combating infectious ... more The identification of novel targets for antimicrobial agents is crucial for combating infectious diseases caused by evolving bacterial pathogens. Components of bacterial toxin–antitoxin (TA) systems have been recognized as promising therapeutic targets. These widespread genetic modules are usually composed of two genes that encode a toxic protein targeting an essential cellular process and an antitoxin that counteracts the activity of the toxin. Uncontrolled toxin expression may elicit a bactericidal effect, so they may be considered “intracellular molecular bombs” that can lead to elimination of their host cells. Based on the molecular nature of antitoxins and their mode of interaction with toxins, TA systems have been classified into six groups. The most prevalent are type II TA systems. Due to their ubiquity among clinical isolates of pathogenic bacteria and the essential processes targeted, they are promising candidates for the development of novel antimicrobial strategies. In t...
Short oligonucleotides binding complementary mRNA essential for bacterial growth could be used as... more Short oligonucleotides binding complementary mRNA essential for bacterial growth could be used as antibacterials provided that they are delivered to bacterial cells. One promising oligonucleotide is peptide nucleic acid (PNA), a synthetic DNA analog with a neutral peptide-like backbone. Unfortunately, bacteria do not import short oligonucleotides, including PNA, from the environment and efficient non-invasive methods for PNA delivery have not been found. To deliver PNA to bacterial cells, we have used vitamin B 12. Most bacteria need to import vitamin B 12 from the environment to sustain life. We have shown that vitamin B 12 covalently linked with PNA, delivers PNA to E. coli cells. Free vitamin B 12 enters E. coli via the TonB-dependent transport system and is recognized by the outer-membrane vitamin B 12-specific BtuB receptor. We engineered the E. coli DbtuB mutant and found that transport of the vitamin B 12-PNA conjugate also requires BtuB. Thus, vitamin B 12 uses the BtuB receptor to deliver PNA through the E. coli outer membrane. Overall, we have proven that vitamin B 12 transport system can be hijacked for PNA delivery to E. coli cells. Further, using Gaussian-force simulated annealing molecular dynamics with umbrella sampling we determined the mechanism of vitamin B 12-PNA permeation through the BtuB receptor protein. BtuB is a beta-barrel occluded by its luminal domain, but we found that partial unfolding of this domain makes the passage of the conjugate mechanically feasible. PNA movement into the beta-barrel is energetically favorable since, while inside BtuB, PNA extends. In addition, we found that BtuB extracellular loops are actively involved in transport through an induced-fit mechanism.
Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature, 2017
Binding of metal ions is an important factor governing the folding and dynamics of RNA. Shielding... more Binding of metal ions is an important factor governing the folding and dynamics of RNA. Shielding of charges in the polyanionic backbone allows RNA to adopt a diverse range of folded structures that give rise to their many functions within the cell. Some RNA sequences fold only in the presence of Mg 2+ , which may be bound via direct interactions or occupy the more diffuse "ion atmosphere" around the RNA. To understand the driving forces for RNA folding, it is important to be able to fully characterize the distribution of metal ions around the RNA. In this work, a combined Grand Canonical Monte Carlo-Molecular Dynamics (GCMC-MD) method is applied to characterize Mg 2+ distributions around folded RNA structures. The GCMC-MD approach identifies known innerand outer-shell Mg 2+ coordination, while also predicting new regions occupied by Mg 2+ that are not observed in crystal structures but that may be relevant in solution, including the case of the Mg 2+ riboswitch, for which alternate Mg 2+ binding sites may have implications for its function. This work represents a significant step forward in establishing a structural and thermodynamic description of RNA−Mg 2+ interactions and their role in RNA structure and function.
Peptide nucleic acids are promising nucleic acid analogs for antisense therapies as they can form... more Peptide nucleic acids are promising nucleic acid analogs for antisense therapies as they can form stable duplex and triplex structures with DNA and RNA. Computational studies of PNA-containing duplexes and triplexes are an important component for guiding their design, yet existing force fields have not been well validated and parameterized with modern computational capabilities. We present updated CHARMM and Amber force fields for PNA that greatly improve the stability of simulated PNAcontaining duplexes and triplexes in comparison with experimental structures and allow such systems to be studied on microsecond time scales. The force field modifications focus on reparametrized PNA backbone torsion angles to match high-level quantum mechanics reference energies for a model compound. The microsecond simulations of PNA-PNA, PNA-DNA, PNA-RNA, and PNA-DNA-PNA complexes also allowed a comprehensive analysis of hydration and ion interactions with such systems.
The widespread emergence of bacterial resistance to existing antibiotics forces the development o... more The widespread emergence of bacterial resistance to existing antibiotics forces the development of new therapeutic agents. The use of short modified oligonucleotides, such as peptide nucleic acids (PNAs), seems a promising strategy. However, the uptake of such oligonucleotides is limited by the bacterial cell wall and is species-dependent. Therefore, new carriers for PNAs should be extensively explored. In this study, we examined the antibacterial activity of vitamin B 12 −PNA conjugates. Vitamin B 12 was covalently linked to a PNA oligomer targeted at the mRNA of an essential acpP gene encoding acyl carrier protein in Escherichia coli. PNA−vitamin B 12 conjugates were synthesized using the Cu(I)-catalyzed 1,3-dipolar cycloaddition. We examined two types of linkers between vitamin B 12 and PNA, including a cleavable disulfide bond. As a positive control for PNA uptake, we used PNA conjugated to the most widely used cell-penetrating peptide (KFF) 3 K. We found that vitamin B 12 −PNA conjugates inhibit E. coli growth at a concentration of 5 μM, similar as (KFF) 3 K−PNA. We also showed that vitamin B 12 −PNA conjugates are stable in the presence of biological media. This study provides the foundation for improving and developing PNAs conjugated to vitamin B 12 as antibacterials.
Clindamycin, a lincosamide antibiotic, binds to 23S ribosomal RNA and inhibits protein synthesis.... more Clindamycin, a lincosamide antibiotic, binds to 23S ribosomal RNA and inhibits protein synthesis. The A2058G mutation in 23S RNA results in bacterial resistance to clindamycin. To understand the influence of this mutation on short-range interactions of clindamycin with 23S RNA, we carried out full-atom molecular dynamics simulations of a ribosome fragment containing clindamycin binding site. We compared the dynamical behavior of this fragment simulated with and without the A2058G mutation. Molecular dynamics simulations suggest that clindamycin in the native ribosomal binding site is more internally flexible than in the A2058G mutant. Only in the native ribosome fragment did we observe intramolecular conformational change of clindamycin around its C7-N1-C10-C11 dihedral. In the mutant, G2058 makes more stable hydrogen bonds with clindamycin hindering its conformational freedom in the ribosome-bound state. Clindamycin binding site is located in the entrance to the tunnel through which the newly synthesized polypeptide leaves the ribosome. We observed that in the native ribosome fragment, clindamycin blocks the passage in the tunnel entrance, whereas in the mutated fragment the aperture is undisturbed due to a different mode of binding of clindamycin in the mutant. Restricted conformational freedom of clindamycin in a position not blocking the tunnel entrance in the A2058G mutant could explain the molecular mechanism of bacterial resistance against clindamycin occurring in this mutant.
Biochimica et Biophysica Acta (BBA) - Biomembranes, 2019
Proteinase 3 (PR3), together with other serine proteases, such as neutrophil elastase (NE) and ca... more Proteinase 3 (PR3), together with other serine proteases, such as neutrophil elastase (NE) and cathepsin G (CG), regulates inflammatory and immune responses. However, in comparison with NE and CG, there is increasing evidence that PR3 functions significantly differ. In particular, PR3 can bind to cell membranes and such membrane-bound PR3 (mbPR3) might be differently involved in the activation of cytokines, growth factors, cellular receptors, and in the regulation of cell apoptosis. For instance, PR3 membrane binding can block some "eat me" signals, notably, phosphatidylserine membrane lipid, and facilitate non-resolving inflammation. Based on the clear evidence that PR3 membrane binding affects the biological functions of PR3, we designed peptidomimetic inhibitors that can remove mbPR3 from the membrane surface in vitro without influencing PR3 catalytic activity. Such inhibitors, which specifically target PR3 binding to membranes, are still lacking. In particular, we found peptidomimetics that inhibit binding of PR3 to POPC:PS liposomes, which mimic the biological environment of PR3.
Journal of Chemical Information and Modeling, 2019
Transport of various molecules facilitated with membrane proteins is necessary for maintaining ho... more Transport of various molecules facilitated with membrane proteins is necessary for maintaining homeostasis in living cells. In humans, dysfunction of these proteins leads to many diseases. Thus, understanding how the membrane proteins function may help using them as therapeutic targets. To successfully investigate the mechanistic aspects of transport, the choice of appropriate methods is crucial. We review the computational methods that have proven most effective in investigating transport events, specifically, deterministic time-dependent classical molecular dynamics and its enhanced sampling variants, as well as methods based on Brownian dynamics. We describe technical aspects of these methods and examples of their novel variants or combinations that have
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Papers by Joanna Trylska