Drug Binding Mode Research Articles

Binding Mode Prediction of Evodiamine within Vanilloid Receptor TRPV1

Accurate assessment of the potential binding mode of drugs is crucial to computer-aided drug design paradigms. It has been reported that evodiamine acts as an agonist of the vanilloid receptor Transient receptor potential vanilloid-1 (TRPV1). However, the precise interaction between evodiamine and TRPV1 was still not fully understood. In this perspective, the homology models of TRPV1 were generated using the crystal structure of the voltage-dependent shaker family K+ channel as a template. We then performed docking and molecular dynamics simulation to gain a better understanding of the probable binding modes of evodiamine within the TRPV1 binding pocket. There are no significant interspecies differences in evodiamine binding in rat, human and rabbit TRPV1 models. Pharmacophore modeling further provided confidence for the validity of the docking studies. This study is the first to shed light on the structural determinants required for the interaction between TRPV1 and evodiamine, and gives new suggestions for the rational design of novel TRPV1 ligands.

Structural Characterization of Binding Mode of Smoking Cessation Drugs to Nicotinic Acetylcholine Receptors through Study of Ligand Complexes with Acetylcholine-binding Protein

Smoking cessation is an important aim in public health worldwide as tobacco smoking causes many preventable deaths. Addiction to tobacco smoking results from the binding of nicotine to nicotinic acetylcholine receptors (nAChRs) in the brain, in particular the α4β2 receptor. One way to aid smoking cessation is by the use of nicotine replacement therapies or partial nAChR agonists like cytisine or varenicline. Here we present the co-crystal structures of cytisine and varenicline in complex with Aplysia californica acetylcholine-binding protein and use these as models to investigate binding of these ligands binding to nAChRs. This analysis of the binding properties of these two partial agonists provides insight into differences with nicotine binding to nAChRs. A mutational analysis reveals that the residues conveying subtype selectivity in nAChRs reside on the binding site complementary face and include features extending beyond the first shell of contacting residues.

Complementary Fluorescence and Phosphorescence Study of the Interaction of Brompheniramine with Human Serum Albumin

Binding of the antihistamine drug brompheniramine (BPA) to human serum albumin (HSA) is studied by measuring quenching of the fluorescence and room temperature phosphorescence (RTP) of tryptophan. The modified Stern-Volmer equation was used to derive association constants and accessible fractions from the steady-state fluorescence data. Decay associated spectra (DAS) revealed three tryptophan fluorescence lifetimes, indicating the presence of three HSA conformations. BPA causes mainly static quenching of the long-living, solvent-exposed conformer. RTP spectra and lifetimes, recorded under deoxygenated conditions in the presence of 0.2 M KI, provided additional kinetic information about the HSA-BPA interactions. Fluorescence DAS that were also recorded in the presence of 0.2 M KI revealed that the solvent-exposed conformer is the major contributor to the RTP signal. The phosphorescence quenching is mostly dynamic at pH 7 and mostly static at pH 9, presumably related to the protonation state of the alkylamino chain of BPA. This provides direct insight into the binding mode of the antihistamine drug, as well as kinetic information at both the nanosecond and the millisecond time scales.

Spectroscopic investigation on the interaction of 3,7-dihydroxyflavone with different isomers of human serum albumin

The interaction mechanism of 3,7-dihydroxyflavone (3,7-diHF) and human serum albumin (HSA) was investigated by fluorescence quenching, fluorescence enhancement, steady-state and time-resolved fluorescence emission and UV–vis absorption spectrometry. The binding site of 3,7-diHF on protein was determined by investigating the spectroscopic properties of 3,7-diHF–HSA complex at pH 7.4 and pH 3.5 individually, and confirmed by the site marker competitive experiments. The binding parameters of 3,7-diHF–HSA complex were estimated by fluorescence quenching experiments, and the data were in good agreement with the results obtained from fluorescence enhancement measurements. A remarkable increase in the fluorescence anisotropy values suggested that 3,7-diHF bound at a motional restricted pocket on HSA. The results indicated that 3,7-diHF, in anionic form, was bound within the hydrophobic pockets of the subdomain IIA of HSA (site I), and stabilised mainly by electrostatic force and ionic interactions. The binding mode of drug–protein was discussed based on above experimental results.

RNA adducts with Na2SeO4 and Na2SeO3 – Stability and structural features

Abstract Selenium compounds are widely available in dietary supplements and have been extensively studied for their antioxidant and anticancer properties. Low blood Se levels were found to be associated with an increased incidence and mortality from various types of cancers. Although many in vivo and clinical trials have been conducted using these compounds, their biochemical and chemical mechanisms of efficacy are the focus of much current research. This study was designed to examine the interaction of Na 2 SeO 4 and Na 2 SeO 3 with RNA in aqueous solution at physiological conditions, using a constant RNA concentration (6.25 mM) and various sodium selenate and sodium selenite/polynucleotide (phosphate) ratios of 1/80, 1/40, 1/20, 1/10, 1/5, 1/2 and 1/1. Fourier transform infrared, UV–Visible spectroscopic methods were used to determine the drug binding modes, the binding constants, and the stability of Na 2 SeO 4 and Na 2 SeO 3 –RNA complexes in aqueous solution. Spectroscopic evidence showed that Na 2 SeO 4 and Na 2 SeO 3 bind to the major and minor grooves of RNA ( via G, A and U bases) with some degree of the Se–phosphate (PO 2 ) interaction for both compounds with overall binding constants of K(Na 2 SeO 4 –RNA) = 8.34 × 10 3 and K(Na 2 SeO 3 –RNA) = 4.57 × 10 3 M −1 . The order of selenium salts–biopolymer stability was Na 2 SeO 4 –RNA > Na 2 SeO 3 –RNA. RNA aggregations occurred at higher selenium concentrations. No biopolymer conformational changes were observed upon Na 2 SeO 4 and Na 2 SeO 3 interactions, while RNA remains in the A-family structure.

Locating the binding sites of anticancer tamoxifen and its metabolites 4-hydroxytamoxifen and endoxifen on bovine serum albumin

The breast anticancer drug tamoxifen and its metabolites bind serum albumins. We located the binding sites of tamoxifen, 4-hydroxytamoxifen and endoxifen on bovine serum albumin (BSA). FTIR, CD and fluorescence spectroscopic methods as well as molecular modeling were used to characterize the drug binding mode, binding constant and the effect of drug binding on BSA stability and conformation. Structural analysis showed that tamoxifen and its metabolites bind BSA via hydrophobic and hydrophilic interactions with overall binding constants of K tam–BSA = 1.96 (±0.2) × 10 4 M −1, K 4-hydroxytam–BSA = 1.80 (±0.4) × 10 4 M −1 and K endox–BSA = 8.01 (±0.8) × 10 3 M −1. The number of bound drug molecules per protein is 1.7 (tamoxifen), 1.4 (4-hydroxitamoxifen) and 1.13 (endoxifen). The participation of several amino acid residues in drug–protein complexes is stabilized by extended hydrogen bonding network with the free binding energy of −13.47 (tamoxifen), −13.79 (4-hydroxtamoxifen) and −12.72 kcal/mol (endoxifen). The order of binding is 4-hydroxy-tamoxen > tamoxifen > endoxifen. BSA conformation was altered by a major reduction of α-helix from 63% (free BSA) to 41% with tamoxifen, to 39% with 4-hydroxytamoxifen, and to 47% with endoxifen. In addition, an increase in turn and random coil structures was found, suggesting partial protein unfolding. These results suggest that serum albumins might act as carrier proteins for tamoxifen and its metabolites in delivering them to target tissues.

Binding of antitumor tamoxifen and its metabolites 4-hydroxytamoxifen and endoxifen to human serum albumin

Tamoxifen is extensively metabolized, and several metabolites have been detected in human serum. The aim of this study was to examine the interaction of human serum albumin (HSA) with tamoxifen and its metabolites 4-hydroxytamoxifen and endoxifen at physiological conditions, using constant protein concentration and various drug contents. FTIR, UV–Visible, CD and fluorescence spectroscopic methods as well as molecular modeling were used to analyse drug binding mode, the binding constant and the effects of drug complexation on HSA stability and conformation. Structural analysis showed that tamoxifen and its metabolites bound HSA via both hydrophobic and hydrophilic interactions with overall binding constants of K tam = 1.8 (±0.2) × 10 4 M −1, K 4-hydroxytam = 1.8 (±0.4) × 10 4 M −1 and K endox = 2.0 (±0.5) × 10 4 M −1. The number of bound drugs per protein is 1.2 (tamoxifen), 1.7 (4-hydroxitamoxifen) and 1.0 (endoxifen). Structural modeling showed the participation of several amino acid residues in drug–HSA complexation, with extended H-bonding network. HSA conformation was altered by tamoxifen and its metabolites with a major reduction of α-helix and an increase in β-sheet, random coil and turn structures, indicating a partial protein unfolding. Our results suggest that serum albumins can act as carrier proteins for tamoxifen and its metabolites in delivering them to target tissues.

Computational study of drug binding to the membrane-bound tetrameric M2 peptide bundle from influenza A virus

The M2 protein of influenza A virus performs the crucial function of transporting protons to the interior of virions enclosed in the endosome. Adamantane drugs, amantadine (AMN) and rimantidine (RMN), block the proton conduction in some strains, and have been used for the treatment and prophylaxis of influenza A infections. The structures of the transmembrane (TM) region of M2 that have been solved in micelles using NMR (residues 23-60) (Schnell and Chou, 2008) and by X-ray crystallography (residues 22-46) (Stouffer et al., 2008) suggest different drug binding sites: external and internal for RMN and AMN, respectively. We have used molecular dynamics (MD) simulations to investigate the nature of the binding site and binding mode of adamantane drugs on the membrane-bound tetrameric M2-TM peptide bundles using as initial conformations the low-pH AMN-bound crystal structure, a high-pH model derived from the drug-free crystal structure, and the high-pH NMR structure. The MD simulations indicate that under both low- and high-pH conditions, AMN is stable inside the tetrameric bundle, spanning the region between residues Val27 to Gly34. At low pH the polar group of AMN is oriented toward the His37 gate, while under high-pH conditions its orientation exhibits large fluctuations. The present MD simulations also suggest that AMN and RMN molecules do not show strong affinity to the external binding sites.

Structural Plasticity and Distinct Drug-Binding Modes of LfrR, a Mycobacterial Efflux Pump Regulator

The TetR-like transcriptional repressor LfrR controls the expression of the gene encoding the Mycobacterium smegmatis efflux pump LfrA, which actively extrudes fluoroquinolones, cationic dyes, and anthracyclines from the cell and promotes intrinsic antibiotic resistance. The crystal structure of the apoprotein form of the repressor reveals a structurally asymmetric homodimer exhibiting local unfolding and a blocked drug-binding site, emphasizing the significant conformational plasticity of the protein necessary for DNA and multidrug recognition. Crystallographic and calorimetric studies of LfrR-drug complexes further confirm the intrinsic flexibility of the homodimer, which provides a dynamic mechanism to broaden multidrug binding specificity and may be a general property of transcriptional repressors regulating microbial efflux pump expression.

A Comparative Study on the Interaction of Cis- and Trans-Platin with DNA and RNA

Cis-diamminedichloroplatinum(II) is a frequently used and very effective chemotherapeutic drug for treatment of various malignancies; however, the trans isomer is clinically ineffective. Cis-platin exerts its antitumor activity by binding to DNA via intrastrand cross-links to d(GpG) (dG = deoxyguanosine) and to d(ApG) (dA = deoxyadenosine), interfering with DNA replication and transcription and causing cell death. The trans-diamminedichloroplatinum(II) isomer also binds DNA, but is clinically ineffective. This study was designed to examine the interactions of cis- and trans-platin with calf thymus DNA and yeast RNA in aqueous solution at physiological conditions, using a constant DNA and RNA concentration (6.25 mM) and various platin salts/polynucleotide (phosphate) ratios of 1/100, 1/50, 1/25, and 1/12.5. Fourier transform infrared, ultraviolet-visible spectroscopic methods were used to determine the drug binding modes, the binding constants, and the stability of cis- and trans-platin-DNA and -RNA complexes in aqueous solution. Spectroscopic evidence showed that cis- and trans-platin bind to the major and minor grooves of DNA (via G, A, T, and C bases), while RNA binding is through G, U, A, and C bases with some degree of the pt-phosphate (PO(2)) interaction for both isomers and overall binding constants of K((cis-platin-DNA)) = 5.51 x 10(4) M(-1), K((trans-platin-DNA)) = 2.26 x 10(4) M(-1), K((cis-platin-RNA)) = 1.9 x 10(4) M(-1), and K((trans-platin-RNA)) = 1.75 x 10(4) M(-1). DNA and RNA aggregations occurred at high platin concentrations. No biopolymer conformational changes were observed upon cis- and trans-platin interactions, while DNA remains in the B-family, and RNA retains its A-family structure. The order of platin compound-polymer stability was cis-platin-DNA > trans-platin-DNA > cis-platin-RNA > trans-platin-RNA.

Inside Cover: Docking Studies of Structurally Diverse Antimalarial Drugs Targeting PfATP6: No Correlation between in silico Binding Affinity and in vitro Antimalarial Activity. (ChemMedChem 9/2009)

The inside cover picture shows the calculated binding modes of antimalarial drugs in the active site of the Plasmodium falciparum calcium ATPase (PfATP6). Docking studies of peroxide-containing antimalarial agents indicate that highly potent compounds have the same affinity for PfATP6 as their biologically inactive derivatives, making the role of PfATP6 inhibition in the mechanism of action of artemisinin doubtful. For more details, see the Full Paper by A. Robert et al. on p. 1469 ff.

Mixed‐Model QSAR at the Glucocorticoid Receptor: Predicting the Binding Mode and Affinity of Psychotropic Drugs

The glucocorticoid receptor (GR) is a member of the nuclear receptor superfamily that affects immune response, development, and metabolism in target tissues. Glucocorticoids are widely used to treat diverse pathophysiological conditions, but their clinical applicability is limited by side effects. A prediction of the binding affinity toward the GR would be beneficial for identifying glucocorticoid-mediated adverse effects triggered by drugs or chemicals. By identifying the binding mode to the GR using flexible docking (software Yeti) and quantifying the binding affinity through multidimensional QSAR (software Quasar), we validated a model family based on 110 compounds, representing four different chemical classes. The correlation with the experimental data (cross-validated r(2)=0.702; predictive r(2)=0.719) suggests that our approach is suited for predicting the binding affinity of related compounds toward the GR. After challenging the model by a series of scramble tests, a consensus approach (software Raptor), and a prediction set, it was incorporated into our VirtualToxLab and used to simulate and quantify the interaction of 24 psychotropic drugs with the GR.

Study of DNA Interactions with Steroidal and Nonsteroidal Estrogen–Platinum (II)–Based Anticancer Drugs

The interactions of the steroidal and nonsteroidal estrogen-platinum (Pt) (II)-based anticancer drugs 16beta-hydroxymethyl-16alpha-[8-(2-pyridin-2-yl-ethylamino)-3,6-dioxaoctyl]-1,3,5(10)-estratrien-3,17betadiol dichloroplatinum (II) (JPM-39), 4-[6-(2'-pyridylethylamino)-butyloxy)-phenyl]-7-methoxy-2,2-dimethyl-3-phenyl-chroman dichloroplatinum (II) (ATG-99), and 1-[(2-aminoethyl)amino]-9,10,10-tris(4-hydroxyphenyl)-9-decene dichloroplatinum (II) (GEB-28) with calf-thymus DNA in vitro using constant DNA concentration and various drug levels were studied. Fourier transform infrared (FTIR) and circular dichroism (CD) were studied with calf-thymus DNA in vitro using constant DNA concentration and various drug levels. FTIR, UV-visible, and CD spectroscopic methods were used to characterize the drug binding mode, the binding constant, and structural variations of DNA in aqueous solution. Spectroscopic evidence showed that the various Pt-based drugs bind indirectly to the major and minor grooves of DNA duplex with some degree of drug-phosphate interaction. The overall binding constants for JPM-39, GEB-28, and ATG-99 are K(JPM-39) = 4.2 (+/-0.75) x 10(3) M(-1), K(GEB-28) = 3.4 (+/-0.65) x 10(3) M(-1), and K(ATG-99) = 2.1 (+/-0.45) x 10(3) M(-1). DNA aggregation occurs at high drug concentration, while DNA remains in the B-family structure.

Study of curcumin and genistein interactions with human serum albumin

Curcumin, the yellow pigment from the rhizoma of Curcuma longa, is a widely studied polyphenolic compound which has a variety of biological activity as anti-inflammatory and antioxidative agent. Genistein one of the flavonoids found in soybean and chickpeas inhibits DNA strand breaks acting as a direct scavenger of reactive oxygen species. Human serum albumin (HSA) with high affinity binding sites is a major transporter for delivering several endogenous compounds and drugs in vivo. The aim of this study was to examine the interactions of curcumin and genistein with human serum albumin at physiological conditions, using constant protein concentration and various pigment contents. FTIR, UV–Visible, CD and fluorescence spectroscopic methods were used to analyse drug binding mode, the binding constant and the effects of pigment complexation on HSA stability and conformation. Structural analysis showed that curcumin and genistein bind HSA via polypeptide polar groups with overall binding constants of K curcumin = 5.5 (±0.8) × 10 4 M −1 and K genistein = 2.4 (±0.40) × 10 4 M −1. The number of bound pigment ( n) is 1.33 for curcumin and 1.49 for genistein. The HSA conformation was altered by pigment complexation with reduction of α-helix and increase of random coil and turn structures suggesting a partial protein unfolding.

Investigation of double‐stranded DNA/drug interaction by ESI/FT ICR: orientation of dissociations relates to stabilizing salt bridges

Noncovalent complexes of DNA and Hoechst 33258 were investigated by ESI-FT/ICR MS in various activation modes (collision-induced dissociation (CID), sustained off-resonance irradiation collision-induced dissociation (SORI-CID), infrared multiphoton dissociation (IRMPD) and electron detachment dissociation (EDD)). The binding selectivity of Hoechst 33258 was confirmed by the comparative study of its noncovalent association with different DNA sequences. The CID spectra of [ds + HO - 5H](5-) obtained with a linear hexapole ion trap resulted in unzipping of the strands. This outcome is a clue to the drug-binding mode, shading light on the localization of the binding sites of Hoechst 33258 to the DNA sequence. The IRMPD and SORI-CID experiments mainly gave DNA backbone cleavages and internal fragment ions. From this result, information on the localization of the binding sites of Hoechst 33258 in the DNA sequence was obtained. No sodium cationization was observed on the DNA sequence ions although they were present on fragmentation of the duplex, indicating that the backbone cleavages were generated from the single strand associated with the Hoechst 33258 where the number of alkali cation is restricted. Under electron detachment (ED) conditions, multiple EDs were achieved for the [ds + HO - 5H](5-) ion without any significant dissociation. The presence of drug appears to enhance the stability of the multiply charged system. It was proposed that the studied noncovalent complex involved the formation of zwitterions and consequently strong salt-bridge interactions between DNA and drug.

Transfer RNA Bindings to Antitumor Estradiol-Platinum(II) Hybrid and Cisplatin

The anticancer platinum (Pt) drugs exert their antitumor activity by direct or indirect Pt-DNA binding. It has been shown that Pt drugs can induce major DNA damage and minor RNA damage during cancer treatment. A recent report showed that a new anticancer estradiol-Pt(II) hybrid molecule (CD-37) binds DNA bases indirectly, while being more effective than cis-diaminedichloroplatinum(II) (cisplatin) against several types of cancer. In this report, we examine the bindings of CD-37 and cisplatin drugs with transfer RNA (tRNA) in vitro and compare the results to those of the corresponding Pt-DNA complexes. Solutions containing various CD-37 or cisplatin concentrations were reacted with tRNA at physiological pH. Using Fourier transform infrared (FTIR), UV-visible, and circular dichroism spectroscopic methods, the drug binding mode, the binding constant, and RNA structural variations are determined for Pt-tRNA complexes in aqueous solution. Structural analysis showed direct binding of cisplatin drug to guanine and adenine N7 sites, while both direct and indirect interactions of CD-37 with tRNA bases and the backbone phosphate group were observed. The overall binding constants estimated were K(CD-37) = 2.77 (+/-0.90) x 10(4) M(1) and K(cisplatin) = 1.72 (+/-0.50) x 10(4) M(1). Major aggregation of tRNA occurs at high CD-37 concentrations, while RNA remains in the A-family structure.

Modeling hERG and its Interactions with Drugs: Recent Advances in Light of Current Potassium Channel Simulations

The hERG K(+) channel is responsible for the rapid delayed rectifier current in cardiac myocytes, and a block of its functioning may be related with the (inherited or drug-induced) long QT syndrome. For this reason, in recent times, some interest has arisen around computational studies aimed at developing hERG/drug models for the prediction of drug binding (docking) modes, in view of the assessment of the hERG blocking potential. On the other hand, voltage-gated K(+) channels have been the subject of molecular simulations for several years, and rigorous protocols for studying the main aspects of their functions (permeation, gating, voltage sensing) have been published. In this article, we briefly introduce these classical computational works on K(+) channels, and then review in depth the reports on the latest advanced modeling studies on hERG. The aim is to put the hERG modeling work in the more general context of the ion channel simulations field, to show the peculiarity of hERG on the one side, and also to indicate some possible new avenues in the use of modeling techniques to increase our knowledge of this important channel.

DNA Interaction with Novel Antitumor Estradiol–Platinum(II) Hybrid Molecule: A Comparative Study with Cisplatin Drug

Platinum(II)-based anticancer drugs are effective for the management and treatment of several types of cancer. Cis-diamminedichloroplatinum(II) (cisplatin) exerts its antitumor activity by binding to DNA via intrastrand cross-links to d(GpG) (dG = deoxyguanosine) and to d(ApG) (dA = deoxyadenosine), causing DNA bending and interfering with DNA replication and transcription. However, the exact binding modes of other platinum(II)-based antitumor drugs to DNA duplex and their mechanism of action have not been clearly investigated. The aim of this study was to examine the binding of a novel anticancer estradiol-platinum(II) hybrid molecule (CD-37) with calf-thymus DNA in vitro and to compare the results with those obtained with cisplatin drug. Solutions containing various CD-37 or cisplatin concentrations were reacted with DNA at physiological pH. Then, using Fourier transform infrared, ultraviolet-visible, and circular dichroism spectroscopic methods, it was possible to characterize the drug binding mode, the binding constant, and structural variations of DNA in aqueous solution. Spectroscopic evidence showed that cisplatin binds to guanine N7 site with minor perturbations of the backbone phosphate group with an overall binding constant of K(cisPt) = 5.73 (+/- 0.45) x 10(4) M(-1). CD-37 binds to DNA duplex via H-bonding network at low drug concentrations with minor perturbations of guanine N7 site at high drug content and with a binding constant of K(CD-37) = 1.0 (+/- 0.15) x 10(4) M(-1). DNA aggregation occurs at high drug concentration, while DNA remains in the B-family structure.

Modeling the hERG potassium channel in a phospholipid bilayer: Molecular dynamics and drug docking studies

The hERG potassium channel has recently been a matter of extensive studies both at experimental and computational levels, because of its possible involvement in the potentially lethal drug-induced long QT syndrome. In this context, in the absence of an experimentally determined 3D structure, to acquire a detailed description of the channel pore and an understanding of atomic determinants for drug binding is of enormous interest at both academic and industrial levels. To contribute to this aim, we first built the open and closed states of the channel by homology modeling techniques, and then submitted both channel models to ns-time-scale MD simulations in explicit membrane. An in-depth analysis of the dynamical behavior of the channel pore with particular attention to the cavity volume was carried out. Finally, using hERG conformations coming from MD simulations, docking experiments were performed to identify a possible binding mode for the most potent hERG channel blocker so far known, the antihistaminic drug astemizole. We show that the combined use of MD and docking is suitable to identify a possible binding mode of drugs in a fairly good agreement with experiments. Moreover, the exploitation of MD snapshots in the docking experiments allowed us to capture some induced-fit effects related to the side chain conformations of Tyr652 and Phe656, which are residues playing a pivotal role in the hERG drug binding.

Interaction of tRNA with Safranal, Crocetin, and Dimethylcrocetin

Saffron is the red dried stigmas of Crocus sativus L. flowers and used both as a spice and as a drug in traditional therapeutic. The biological activity of saffron in modern medicine is in development. Its numerous applications as an anti-oxidant and anti-cancer agent are due to its secondary metabolites and their derivatives (safranal, crocins, crocetin, dimethylcrocetin). The aim of this study was to examine the interaction of transfer RNA with safranal, crocetin, and dimethylcrocetin in aqueous solution at physiological conditions. Constant tRNA concentration (6.25 mM) and various drug/tRNA (phosphate) molar ratios of 1/48 to 1/8 were used. FT-IR and UV-Visible difference spectroscopic methods have been applied to determine the drug binding mode, the binding constants and the effects of drug complexation on the stability and conformation of tRNA duplex. External binding mode was observed for safranal crocetin and dimethylcrocetin, with overall binding constants Ksafranal = 6.8 (± 0.34) × 103 M−1, KCRT = 1.4 (± 0.31) × 104 M−1, and KDMCRT = 3.4 (± 0.30) × 104 M−1. Transfer RNA remains in the A-family structure, upon safranal, crocetin and dimethylcrocetin complexation.