Major Binding Mode Research Articles

Unveiling the multifaceted interactions of antitumor drug mitoxantrone with ct-DNA through biophysical and in silico studies

Mitoxantrone, an anthraquinone derivative, is a widely used anticancer drug with its well-known ability to engage in complex interactions with DNA. Although known for its intercalating ability, the enigma surrounding its binding modes with DNA persists. The existing corpus of literature primarily focuses on mitoxantrone-DNA interactions with short DNA sequences, thereby yielding insights into its interactive nature is limited to this specific sequence. This study aims to elucidate the diverse modes with which mitoxantrone interacts with calf thymus DNA using a combination of spectroscopy, calorimetry and in silico studies. The findings from spectroscopic, calorimetric and molecular dynamic results in correlation with existing literature, unveil a fascinating narrative: mitoxantrone intercalates at lower concentrations but promotes condensation at higher concentrations. Although intercalation with side chains positioned in the minor/major groove is the major binding mode in GC-rich sequences, molecular modelling studies hint at an alternative binding mode in AT-rich sequences where it exclusively displays pure electrostatic interaction. These findings underscore the pivotal role of both drug structure and base sequence in dictating binding mode and affinity. Such insights not only deepen the understanding of structure-activity relationships but also hold promise for guiding future drug design strategies.

A second life for the crystallographic structure of Berenil-dodecanucleotide complex: a computational revisitation thirty years after its publication

AbstractThis study revisits the pioneering work of Professor Neidle, and co-workers, on the crystal structure of complexes formed between groove binders and DNA sequences. The original research revealed a DNA-ligand complex consisting of a dodecanucleotide bound with Berenil [1,3-bis(4′-amidinophenyl)-triazene] an anti-trypanocidal drug. This article aims to delve deeper into the structural dynamics of this system, showcasing the role played by water molecules in stabilizing the interaction between the ligand and the DNA. With this work, we reevaluate the findings from the original crystallographic study by employing modern molecular dynamics techniques, including Supervised Molecular Dynamics (SuMD) for generating binding trajectories, Thermal Titration Molecular Dynamics for assessing unbinding events, and AquaMMapS to identify regions occupied by stationary water molecules. The study addresses a minor and a major groove binding mode and assesses their strength and specificity using TTMD simulations, generating unbinding trajectories. This comprehensive approach integrates the understanding of the interaction of this DNA-ligand complex, which originated with the valuable work of Professor Neidle, resulting in an in-depth insight into the pivotal role of water molecules with this DNA, a behavior detected and extendable even to other nucleic acid complexes.

Phytochemical analysis and pharmacological activities of essential oils extracted from Zingiber officinale (Roscoe) used in mediterranean diet: in vitro and in silico studies

ABSTRACT For their pungency and tanginess, ginger rhizomes (Zingiber officinale) are often used in cooking. Ginger has long been used in traditional medicine and home remedies to treat pain and inflammation. This research tests Z. officinale (EOZ) essential oils’ chemical composition, insecticidal, and antioxidant properties in vitro and in silico. EOZ yielded 0.69% of root mass with 23 chemicals. The main chemicals in EOZ were α-zingiberene (23.850%), Geranial (14.160%), and (E,E)-α-farnesene (9.980%). EOZ demonstrated noteworthy antioxidant activity in all tests, with an IC50 of 9.53 ± 0.41 mg/mL in the DPPH test and an EC50 of 87.46 ± 3.19 mg/mL in the FRAP system. The results indicate that EOZ has strong efficacy against C. maculatus, even at low concentrations (1.00 μL/100 g). Note that EOZ killed 20 ± 0% and 13.33 ± 4.44% of adult C. maculatus in inhalation and contact tests, respectively. High concentration (20.00 μL/100 g) resulted in 100% adult mortality in inhalation tests and 96.67 ± 4.44% mortality in contact testing. We then used molecular docking to identify EO major component binding modes by targeting insecticidal and antioxidant protein structures 1R20 and 1R4U. This research helps create insecticides and antioxidants from common ginger essential oil.

Complexed Photosensitizer of Hypericin with G-Quadruplex: Structure-Dependent Behavior.

Despite the increased interest of visible-light-absorbing compound Hypericin (Hyp) in photodiagnosis, photocatalysis, and photodynamic therapy (PDT) applications, a major obstacle still exists; i.e., the photoactivity is diminished due to the facile aggregation of Hyp in aqueous environment that induces excited-state quenching. Herein, we explore the excited-state property of Hyp bound to the DNA G-quadruplex by combining multiple steady-state and time-resolved spectroscopy. We find that the aggregation-induced quenching effect can be successfully prevented by appropriate G-quadruplex binders that disperse Hyp into monomer. The binding of Hyp/G-quadruplex is selective, however, exhibiting a preferential binding toward parallel G-quadruplexes (c-kit2, C14B1, STAT3, S50, and PS2.M), over antiparallel or hybrid G-quadruplex (Tel22, TBA). The excited-state property of Hyp is highly related to the binding behavior, showing a consistent trend that the better the Hyp/G-quadruplex binding, the longer the triplet 3Hyp* lifetime and the higher the efficiency to produce 1O2. For Hyp/c-kit2, the major binding mode is 5'-end stacking, which offers protection from collisional quenching reactions and ensures a stable photocycle of 3Hyp*-O2 energy transfer forming 1O2, leading to the highest 1O2 quantum yield (0.67) with superior photostability. These findings open possibilities of developing Hyp/G-quadruplex complex as a biocompatible photosensitizer for PDT applications, etc.

Biosynthesis of Vanillin by Rational Design of Enoyl-CoA Hydratase/Lyase.

Vanillin holds significant importance as a flavoring agent in various industries, including food, pharmaceuticals, and cosmetics. The CoA-dependent pathway for the biosynthesis of vanillin from ferulic acid involved feruloyl-CoA synthase (Fcs) and enoyl-CoA hydratase/lyase (Ech). In this research, the Fcs and Ech were derived from Streptomyces sp. strain V-1. The sequence conservation and structural features of Ech were analyzed by computational techniques including sequence alignment and molecular dynamics simulation. After detailed study for the major binding modes and key amino acid residues between Ech and substrates, a series of mutations (F74W, A130G, A130G/T132S, R147Q, Q255R, ΔT90, ΔTGPEIL, ΔN1-11, ΔC260-287) were obtained by rational design. Finally, the yield of vanillin produced by these mutants was verified by whole-cell catalysis. The results indicated that three mutants, F74W, Q147R, and ΔN1-11, showed higher yields than wild-type Ech. Molecular dynamics simulations and residue energy decomposition identified the basic residues K37, R38, K561, and R564 as the key residues affecting the free energy of binding between Ech and feruloyl-coenzyme A (FCA). The large changes in electrostatic interacting and polar solvating energies caused by the mutations may lead to decreased enzyme activity. This study provides important theoretical guidance as well as experimental data for the biosynthetic pathway of vanillin.

An ensemble of cadherin-catenin-vinculin complex employs vinculin as the major F-actin binding mode

The cell-cell adhesion cadherin-catenin complexes recruit vinculin to the adherens junction (AJ) to modulate the mechanical couplings between neighboring cells. However, it is unclear how vinculin influences the AJ structure and function. Here, we identified two patches of salt bridges that lock vinculin in the head-tail autoinhibited conformation and reconstituted the full-length vinculin activation mimetics bound to the cadherin-catenin complex. The cadherin-catenin-vinculin complex contains multiple disordered linkers and is highly dynamic, which poses a challenge for structural studies. We determined the ensemble conformation of this complex using small-angle x-ray and selective deuteration/contrast variation small-angle neutron scattering. In the complex, both α-catenin and vinculin adopt an ensemble of flexible conformations, but vinculin has fully open conformations with the vinculin head and actin-binding tail domains well separated from each other. F-actin binding experiments show that the cadherin-catenin-vinculin complex binds and bundles F-actin. However, when the vinculin actin-binding domain is removed from the complex, only a minor fraction of the complex binds to F-actin. The results show that the dynamic cadherin-catenin-vinculin complex employs vinculin as the primary F-actin binding mode to strengthen AJ-cytoskeleton interactions.

Peering into the future of SARS-CoV-2 evolution using a computational model of the SARS-CoV-2 serum antibody repertoire

Abstract The global emergence of SARS-CoV-2 has provided a wealth of immunological data, including more than 600 solved structures of SARS-CoV-2 Spike antibodies (Abs) available in the Protein Data Bank. We reasoned that this data could be used to generate a computational model of the serum Ab repertoire. We extracted a set of 275 non-redundant Abs, and clustered them using an unsupervised clustering method based on their contact positions with Spike. We identified 9 unique Ab clusters that together summarize the major binding modes of Abs to the Spike protein. To characterize each cluster, we mined 65 published papers extracting detailed information on each Ab. We then demonstrated Spike Ab contacts could be predicted based on the Ab’s CDRs sequence. Next, we developed a computational method for predicting immune escape. We extracted the set of all Spike mutations in SARS-CoV-2 sequences from GISAID. To assess the effects of single mutations on immune escape, we computed the ΔΔG for each Ab-mutation pair and constructed an aggregated escape score for each mutation. Our predicted escape scores were positively correlated with published experimental measurements of Ab escape. We used our escape score to rank novel viral variants and are dynamically ranking strains and novel variants of interest as they appear. To further validate its predictive power, we chose a set of predicted Ab-escaping mutations and are testing their escape from Abs in human sera samples. Taken together, our analysis demonstrates the feasibility of generating a computational model of the serum Ab repertoire, which may be used for the rapid characterization of novel Abs, and to computationally estimate the effects of mutations on immune escape from Ab recognition. Supposed by the Israel Science Foundation (ISF) grant no. 2683/21, and the NIH\NIAID CEIRR under contract 75N9302100016, St Jude Children’s Research Hospital.

Single crystal structure of nitro terminated Azo Schiff base: DNA binding, antioxidant, enzyme inhibitory and photo-isomerization investigation

Novel azo schiff base compound, namely, 2‑hydroxy-5-[(E)-phenyldiazenyl] benzaldehyde (AzNP) was synthesized by reacting 2‑hydroxy-5-[(E)-phenyldiazenyl]benzaldehyde and 4-(4-nitrophenoxy)benzenamine under inert conditions. The synthesized compound AzNP was characterized by Fourier transform (FT-IR), Nuclear magnetic resonance (NMR) and Ultraviolet-visible (UV–vis) spectroscopy. The solid state structure was elucidated by single-crystal X-ray crystallography. The compound was crystallized in the Monoclinic system with P 1 21/c1 space group, having a = 6.4859(3 Å, b = 15.6673(7) Å, c = 19.7908(9) Å, β = 90.945(2), and Z = 4, showing only one intermolecular C21-H21...N2 interaction, with donor and acceptor distance of 3.5774(16) Å, responsible for the elegant 3D network of the structure. DFT calculations were performed for structure-property relationship information of AzNP. In order to comprehend the selectivity behavior of the compound, binding interaction with CT-DNA was examined using standard optical spectroscopy. The observed hypochromism in the binding studies indicated that ligand substantially interact via the groove binding mode (Kb =9.91×103 mol−1dm3) with the DNA. Further assessment of binding mode of AzNP with DNA was made through hydrodynamic measurement and molecular docking analysis which confirmed the major groove binding mode of the ligand. Additionally, the DPPH (2,2-diphenyl-1-picrylhydrazyl), the ferric reducing power, and the α-amylase inhibition assays were used to evaluate the biological significance. Analysis of Cis-Trans conformational changes was also done to investigate photoisomerization properties of synthesized compound AzNP.

Molecular interplay of ADAMTS13-MDTCS and von willebrand Factor-A2: deepened insights from extensive atomistic simulations

Thrombotic thrombocytopenic purpura (TTP) is a rare and life-threatening disease. One hallmark is severe ADAMTS13 deficiency, causing ultra-large von Willebrand factor (VWF) multimers to accumulate, leading to microthrombi and lastly to microangiopathic hemolytic anemia and severe thrombocytopenia. Despite great success in recent decades, the molecular picture of the interaction between VWF and ADAMTS13 remains vague. Here, we utilized modern replica-exchange molecular dynamics simulations with the TIGER2h method to sample a vast configurational space of the isolated ADAMTS13-MDTCS domains and the exposure to its substrate and activating cofactor – the unraveled VWF-A2 domain. The sampling of binding sites and conformations was guided and filtered in agreement with available experimental evidence. We provide comprehensive information on exosites for each domain and direct pairs of interacting amino acids, for the first time. The major binding cluster for the active site of the MP domain contrasts the previous mapping of VWF-A2 residues and reciprocal binding pockets. Two major binding modes are revealed and provide access to conformational changes of an extended gatekeeper tetrad upon overcoming local latency during substrate binding and to a dedicated recruitment mechanism. Our work adds the first molecular interaction model that places previous experimental results in perspective to better understand disease-related mutations towards improved therapies. Numerous empirical targets are proposed to verify the given binding modes, to refine the overall picture of MP binding pockets, the role of Dis binding in MP activation and the passage of the Cys-rich domain through VWF-A2, thus deepening the understanding of a highly dynamic interplay. Communicated by Ramaswamy H. Sarma

Exploring the interaction of guanidine ligands Amiloride, Rimeporide and Cariporide with DNA for understanding their role as inhibitors of Na+/H+ exchangers (NHEs): A spectroscopic and molecular docking investigation

The inhibition of Na+/H+ Exchangers (NHEs) has shown efficacy in the pathology of several diseases like tumors, cardiovascular, and neurological disorders. The role of guanidine ligands such as amiloride, cariporide, and rimeporide as NHE inhibitors is very well documented but their interaction studies with genomic DNA are still unexplored. In this study, a combination of various biophysical and molecular docking studies was employed to investigate their binding aspects.UV–Visible, fluorescence, and circular dichroism (CD) studies indicated that guanidine ligands bind to the grooves of Calf Thymus DNA (ctDNA). Fluorescence titration studies depict that amiloride binds to ctDNA with a binding constant in the order of 102 M−1 and free energy change (ΔG0) of −14.05 KJ mol−1. Competitive fluorescence studies indicated the minor groove binding property of amiloride, whereas major groove binding mode was deduced for rimeporide and cariporide. Molecular docking studies were also found to be in accordance with the experimental results, revealing the information about the binding energy of the guanidine ligand-ctDNA complex. The docked structures depicted binding energy of −6.4 kcal mol−1 for amiloride and − 6.6 kcal mol−1 for rimeporide and cariporide. Such physicochemical studies of DNA-ligand interactions may facilitate the understanding of the mechanisms of NHE inhibition.

NASP maintains histone H3–H4 homeostasis through two distinct H3 binding modes

Histone chaperones regulate all aspects of histone metabolism. NASP is a major histone chaperone for H3–H4 dimers critical for preventing histone degradation. Here, we identify two distinct histone binding modes of NASP and reveal how they cooperate to ensure histone H3–H4 supply. We determine the structures of a sNASP dimer, a complex of a sNASP dimer with two H3 α3 peptides, and the sNASP–H3–H4–ASF1b co-chaperone complex. This captures distinct functionalities of NASP and identifies two distinct binding modes involving the H3 α3 helix and the H3 αN region, respectively. Functional studies demonstrate the H3 αN-interaction represents the major binding mode of NASP in cells and shielding of the H3 αN region by NASP is essential in maintaining the H3–H4 histone soluble pool. In conclusion, our studies uncover the molecular basis of NASP as a major H3–H4 chaperone in guarding histone homeostasis.

Dynamic and Thermodynamic Investigation on the Interaction of Bovine Serum Albumin with an Anticancer Pt Complex Containing Dithiocarbamate Using Molecular Docking and Spectroscopic Methods

Cisplatin is used for human cancers treatment, such as bladder and ovarian, endometrial and cervical, head and neck, and testicular cancer, but its clinical use has been limited by some serious side effects. Therefore S,S donor ligands such as dithiocarbamates can be used to prepare anticancer Pt-drugs and reduce some side effects. The purpose of this study is the interaction ability of designed Pt complex, [Pt(bpy)(tertamyl.dtc)]NO3, where bpy is bipyridine and tertamyl-dtc is (2-methyl butan-dithiocarbamate), with bovine serum albumin (BSA) using fluorescence emission titration, UV-Vis spectroscopy, and molecular docking methods. The obtained negative values of enthalpy, ΔH 0 b, and entropy, ΔS 0 b, showed that a major binding mode between Pt complex and BSA is the van der Waals and hydrogen bond. The negative value of Gibbs free energy achieved via the UV-Vis technique also exhibited a spontaneous interaction. In addition, the results related to emission titration displayed that the fluorescence quenching of protein by Pt complex is a static quenching mechanism. Moreover, the binding parameters, including apparent biomolecular quenching constant (k q), the number of binding sites (n), and binding constant (K b) were obtained by the Stern–Volmer equation. These values of K b, k q, and n were also obtained as 3 × 108 M−1, 3 × 1014 M−1 S−1, and 1.4, respectively. By the use of spectroscopic kinetics model, this interaction was studied and the result showed a second-order type. Finally, the binding of Pt complexes to BSA was verified by the molecular docking method. The binding free energy and K i for the Pt compounds were obtained as −6.6 kcal mol−1 and 14.48, respectively. The results of molecular docking demonstrate the position of binding of Pt complex on BSA is the site I in the subdomain IIA.

Deep structural insights into RNA-binding disordered protein regions.

Recent efforts to identify RNA binding proteins in various organisms and cellular contexts have yielded a large collection of proteins that are capable of RNA binding in the absence of conventional RNA recognition domains. Many of the recently identified RNA interaction motifs fall into intrinsically disordered protein regions (IDRs). While the recognition mode and specificity of globular RNA binding elements have been thoroughly investigated and described, much less is known about the way IDRs can recognize their RNA partners. Our aim was to summarize the current state of structural knowledge on the RNA binding modes of disordered protein regions and to propose a classification system based on their sequential and structural properties. Through a detailed structural analysis of the complexes that contain disordered protein regions binding to RNA, we found two major binding modes that represent different recognition strategies and, most likely, functions. We compared these examples with DNA binding disordered proteins and found key differences stemming from the nucleic acids as well as similar binding strategies, implying a broader substrate acceptance by these proteins. Due to the very limited number of known structures, we integrated molecular dynamics simulations in our study, whose results support the proposed structural preferences of specific RNA‐binding IDRs. To broaden the scope of our review, we included a brief analysis of RNA‐binding small molecules and compared their structural characteristics and RNA recognition strategies to the RNA‐binding IDRs.This article is categorized under:RNA Structure and Dynamics > RNA Structure, Dynamics, and ChemistryRNA Interactions with Proteins and Other Molecules > Protein–RNA RecognitionRNA Interactions with Proteins and Other Molecules > Small Molecule–RNA Interactions

Regulation of MLL1 Methyltransferase Activity in Two Distinct Nucleosome Binding Modes.

Cryo-EM structures of the KMT2A/MLL1 core complex bound on nucleosome core particles (NCPs) suggest unusual rotational dynamics of the MLL1 complex approaching its physiological substrate. However, the functional implication of such dynamics remains unclear. Here, we show that the MLL1 core complex also shows high rotational dynamics bound on the NCP carrying the catalytically inert histone H3 lysine 4 to methionine (K4M) mutation. There are two major binding modes of the MLL1 complex on the NCPK4M. Importantly, disruption of only one of the binding modes compromised the overall MLL1 activity in an NCP-specific manner. We propose that the MLL1 core complex probably exists in an equilibrium of poised and active binding modes. The high rotational dynamics of the MLL1 complex on the NCP is a feature that can be exploited for loci-specific regulation of H3K4 methylation in higher eukaryotes.

Reexamination of N-terminal domains of syntaxin-1 in vesicle fusion from central murine synapses.

Syntaxin-1 (STX1) and Munc18-1 are two requisite components of synaptic vesicular release machinery, so much so synaptic transmission cannot proceed in their absence. They form a tight complex through two major binding modes: through STX1's N-peptide and through STX1's closed conformation driven by its Habc- domain. However, physiological roles of these two reportedly different binding modes in synapses are still controversial. Here we characterized the roles of STX1's N-peptide, Habc-domain, and open conformation with and without N-peptide deletion using our STX1-null mouse model system and exogenous reintroduction of STX1A mutants. We show, on the contrary to the general view, that the Habc-domain is absolutely required and N-peptide is dispensable for synaptic transmission. However, STX1A's N-peptide plays a regulatory role, particularly in the Ca2+-sensitivity and the short-term plasticity of vesicular release, whereas STX1's open conformation governs the vesicle fusogenicity. Strikingly, we also show neurotransmitter release still proceeds when the two interaction modes between STX1A and Munc18-1 are presumably intervened, necessitating a refinement of the conceptualization of STX1A-Munc18-1 interaction.

Accuracy and precision of binding free energy prediction for a tacrine related lead inhibitor of acetylcholinesterase with an arsenal of supercomputerized molecular modelling methods: a comparative study

Nowadays, advanced computational chemistry methods offer various strategies for revealing prospective hit structures in drug development essentially through accurate binding free energy predictions. After the era of molecular docking and quantitative structure-activity relationships, much interest has been lately oriented to perturbed molecular dynamic approaches like replica exchange with solute tempering and free energy perturbation (REST/FEP) and the potential of the mean force with adaptive biasing and accelerated weight histograms (PMF/AWH). Both of these receptor-based techniques can exploit exascale CPU&GPU supercomputers to achieve high throughput performance. In this fundamental study, we have compared the predictive power of a panel of supercomputerized molecular modelling methods to distinguish the major binding modes and the corresponding binding free energies of a promising tacrine related potential antialzheimerics in human acetylcholinesterase. The binding free energies were estimated using flexible molecular docking, molecular mechanics/generalized Born surface area/Poisson-Boltzmann surface area (MM/GBSA/PBSA), transmutation REST/FEP with 12 x 5 ns/λ windows, annihilation FEP with 20 x 5 ns/λ steps, PMF with weight histogram analysis method (WHAM) and 40 x 5 ns samples, and PMF/AWH with 10 x 100 ns replicas. Confrontation of the classical approaches such as canonical molecular dynamics and molecular docking with alchemical calculations and steered molecular dynamics enabled us to show how large errors in ΔG predictions can be expected if these in silico methods are employed in the elucidation of a common case of enzyme inhibition. Communicated by Ramaswamy H. Sarma

Thermodynamic Evaluation of the Interactions between Anticancer Pt(II) Complexes and Model Proteins.

In this work, we have analysed the binding of the Pt(II) complexes ([PtCl(4′-phenyl-2,2′:6′,2″-terpyridine)](CF3SO3) (1), [PtI(4′-phenyl-2,2′:6′,2″-terpyridine)](CF3SO3) (2) and [PtCl(1,3-di(2-pyridyl)benzene) (3)] with selected model proteins (hen egg-white lysozyme, HEWL, and ribonuclease A, RNase A). Platinum coordination compounds are intensively studied to develop improved anticancer agents. In this regard, a critical issue is the possible role of Pt-protein interactions in their mechanisms of action. Multiple techniques such as differential scanning calorimetry (DSC), electrospray ionization mass spectrometry (ESI-MS) and UV-Vis absorbance titrations were used to enlighten the details of the binding to the different biosubstrates. On the one hand, it may be concluded that the affinity of 3 for the proteins is low. On the other hand, 1 and 2 strongly bind them, but with major binding mode differences when switching from HEWL to RNase A. Both 1 and 2 bind to HEWL with a non-specific (DSC) and non-covalent (ESI-MS) binding mode, dominated by a 1:1 binding stoichiometry (UV-Vis). ESI-MS data indicate a protein-driven chloride loss that does not convert into a covalent bond, likely due to the unfavourable complexes’ geometries and steric hindrance. This result, together with the significant changes of the absorbance profiles of the complex upon interaction, suggest an electrostatic binding mode supported by some stacking interaction of the aromatic ligand. Very differently, in the case of RNase A, slow formation of covalent adducts occurs (DSC, ESI-MS). The reactivity is higher for the iodo-compound 2, in agreement with iodine lability higher than chlorine.

Elucidation of binding interactions and mechanism of Fludarabine with dsDNA via multispectroscopic and molecular docking studies

Fludarabine is a purine derivative, anti-neoplastic drug and is still being used in the treatments of chronic lymphocytic leukemia, small lymphocytic lymphoma, acute myeloid leukemia, Non-Hodgkin’s lymphoma. It achieves its function by interacting with DNA. Therefore, the binding interactions of such drugs with deoxyribonucleic acid (DNA) is an important subject for pharmaceutical and biochemical studies aiming at designing better DNA binding drugs. Although DNA binding mode of some of the anti-neoplastic drugs has been studied, DNA interaction of Fludarabine has not been explored yet. For this reason, this work has been dedicated to deciphering the experimental and theoretical investigation of Fludarabine binding mechanism via multispectroscopic techniques including UV absorption spectroscopy, thermal denaturation, fluorescence and FTIR spectroscopy, electrochemical and viscosity measurement methods as well as with molecular docking studies under physiological conditions. We observed in the lowest energy docking poses that Fludarabine binds to DNA via major groove binding mode. The nonplanar and extended structure and hydrogen bonding interactions of Fludarabine with the Adenine-Thymine base-pair played a very decisive role in the binding mode as supported by the experimental results.

Study on the interactions between B-norcholesteryl benzimidazole compounds with ct-DNA.

The study of molecule-DNA interaction is very important for designing an improved therapeutic agent. In previous studies, we synthesized some B-norcholesteryl benzimidazole compounds, and the tests on cancer cells showed that these compounds had good invitro anti-cancer activities. In order to further investigate mechanism of their actions, three different B-norcholesteryl benzimidazole compounds were selected and interaction of these compounds with the calf thymus DNA (ct-DNA) was monitored by using various methods including UV-Vis and fluorescence spectroscopic techniques, viscosity measurement, and circular dichroism (CD). The results proved a hypochromic effect accompanied with a slight red-shift due to the interaction of the molecules with ct-DNA. According to the UV-Vis and fluorescence spectra, the mentioned compounds were bound to DNA, preferentially through partial intercalation into the DNA helix. Moreover, the ethidium bromide (EB) and Hoechst 33258 competitive binding experiments were also used to confirm the interaction mode of the compounds with ct-DNA. In the Hoechst 33258 displacement experiment, no significant change in the fluorescence intensity was observed. Additional assays such as iodide quenching, viscosity, and CD spectroscopy further confirmed that intercalation should be the major binding mode of the selected compounds with DNA. The cytotoxicity of these three compounds was also evaluated by MTT method, and the results confirmed that binding ability of these compounds to DNA was consistent with their cytotoxicity behavior. The experimental results indicated a higher binding affinity for compound 3 compared to the other compounds. This research provided a better understanding on the molecular mechanism of the interaction between B-norcholesteryl benzimidazole compounds and tumor cells, and offered a beneficial perspective to the designation of novel B-norsteroidal anticancer compounds.

Complexed crystal structure of SSB reveals a novel single-stranded DNA binding mode (SSB)3:1: Phe60 is not crucial for defining binding paths

Single-stranded DNA-binding protein (SSB) is essential to cells as it participates in DNA metabolic processes, such as DNA replication, repair, and recombination. Escherichia coli SSB (EcSSB) tetramer cooperatively binds and wraps ssDNA in two major binding modes. In this study, we report the complex crystal structure of Pseudomonas aeruginosa SSB (PaSSB) with ssDNA dT20 at 2.39 Å resolution (PDB entry 6JDG) that revealed a new binding mode, namely, (SSB)3:1. In the (SSB)65 mode revealed by the EcSSB–dC35 complex structure, all four subunits fully participate in the binding to ssDNA. However, only three subunits in the PaSSB tetramer can participate in wrapping ssDNA in the (SSB)3:1 mode. The bound ssDNA in the PaSSB–ssDNA complex adopts an Ω-shaped conformation rather than a χ-shaped conformation in the (SSB)65 mode possibly due to the disability of Phe60. Phe60 is known to play a critical role in defining DNA-binding paths and promoting the wrapping of ssDNA around SSB tetramers. However, it is not important in the (SSB)3:1 mode. The ssDNA binding path revealed by our structural evidence suggests that ssDNA occupies half of the binding sites of the two subunits and slightly comes into contact with the ssDNA binding sites of the third subunit. Accordingly, we propose and sketch a possible wrapping mechanism of SSB via this novel ssDNA-binding mode, (SSB)3:1.