Insulin-like effects of polyamines spermine binding to fat cells and fat cell membranes

Two naturally occurring polyamines, spermine and spermidine, mimic the action of insulin on lipid and glucose metabolism in adipocytes. To evaluate the role of cell membranes in the action of polyamines, studies of [¹⁴C] spermine binding using an oil separation method were conducted in isolated rat adipocytes and adipose cell membranes. Spermine binding and dissociation in fat cells and fat cell membranes were rapid and complete within 3–6 min. Following a 30-min incubation of [¹⁴C] spermine with fat cell membranes, over 90% of bound [¹⁴C] spermine was dissociable while under similar conditions only 25% of bound [¹⁴C] spermine was dissociable in cells. The non-dissociable fractions in cells likely represented intracellular accumulation. Binding and stimulation of glucose oxidation were demonstrated at similar concentrations. Bound spermine was displaced by spermine, spermidine and 1,8-diaminooctane with greater efficacy than putrescine (a polyamine devoid of insulin-like properties) or insulin. Similarly, polyamines did not complete with insulin for binding to isolated adipocytes. It appears, therefore, that polyamines initiate their insulin-like effects by interacting with the cell membrane at sites which are common to biologically active polyamines and which are distinct from the insulin receptor.     

Nuclear magnetic resonance structure of calcium-binding protein 1 in a Ca(2+) -bound closed state: Implications for target recognition

Calcium‐binding protein 1 (CaBP1), a neuron‐specific member of the calmodulin (CaM) superfamily, regulates the Ca²⁺‐dependent activity of inositol 1,4,5‐triphosphate receptors (InsP3Rs) and various voltage‐gated Ca²⁺ channels. Here, we present the NMR structure of full‐length CaBP1 with Ca²⁺ bound at the first, third, and fourth EF‐hands. A total of 1250 nuclear Overhauser effect distance measurements and 70 residual dipolar coupling restraints define the overall main chain structure with a root‐mean‐squared deviation of 0.54 Å (N‐domain) and 0.48 Å (C‐domain). The first 18 residues from the N‐terminus in CaBP1 (located upstream of the first EF‐hand) are structurally disordered and solvent exposed. The Ca²⁺‐saturated CaBP1 structure contains two independent domains separated by a flexible central linker similar to that in calmodulin and troponin C. The N‐domain structure of CaBP1 contains two EF‐hands (EF1 and EF2), both in a closed conformation [interhelical angles = 129° (EF1) and 142° (EF2)]. The C‐domain contains EF3 and EF4 in the familiar Ca²⁺‐bound open conformation [interhelical angles = 105° (EF3) and 91° (EF4)]. Surprisingly, the N‐domain adopts the same closed conformation in the presence or absence of Ca²⁺ bound at EF1. The Ca²⁺‐bound closed conformation of EF1 is reminiscent of Ca²⁺‐bound EF‐hands in a closed conformation found in cardiac troponin C and calpain. We propose that the Ca²⁺‐bound closed conformation of EF1 in CaBP1 might undergo an induced‐fit opening only in the presence of a specific target protein, and thus may help explain the highly specialized target binding by CaBP1.     

A molecular dynamics simulation of the flavin mononucleotide–RNA aptamer complex

We report on an unrestrained molecular dynamics simulation of the flavin mononucleotide (FMN)–RNA aptamer. The simulated average structure maintains both cross‐strand and intermolecular FMN–RNA nuclear Overhauser effects from the nmr experiments and has all qualitative features of the nmr structure including the G10–U12–A25 base triple and the A13–G24, A8–G28, and G9–G27 mismatches. However, the relative orientation of the hairpin loop to the remaining part of the molecule differs from the nmr structure. The simulation predicts that the flexible phosphoglycerol part of FMN moves toward G27 and forms hydrogen bonds. There are structurally long‐lived water molecules in the FMN binding pocket forming hydrogen bonds within FMN and between FMN and RNA. In addition, long‐lived water is found bridging primarily RNA backbone atoms. A general feature of the environment of long‐lived “structural” water is at least two and in most cases three or four potential acceptor atoms. The 2′‐OH group of RNA usually acts as an acceptor in interactions with the solvent. There are almost no intrastrand O2′H(n)⋮O4′(n + 1) hydrogen bonds within the RNA backbone. In the standard case the preferred orientation of the 2′‐OH hydrogen atoms is approximately toward O3′ of the same nucleotide. However, a relatively large number of conformations with the backbone torsional angle γ in the trans orientation is found. A survey of all experimental RNA x‐ray structures shows that this backbone conformation occurs but is less frequent than found in the simulation. Experimental nmr RNA aptamer structures have a higher fraction of this conformation as compared to the x‐ray structures. The backbone conformation of nucleotide n + 1 with the torsional angle γ in the trans orientation leads to a relatively short distance between 2′‐OH(n) and O5′(n + 1), enabling hydrogen‐bond formation. In this case the preferred orientation of the 2′‐OH hydrogen atom is approximately toward O5′(n + 1). We find two relatively short and dynamically stable types of backbone–backbone next‐neighbor contacts, namely C2′(H)(n)⋮O4′(n + 1) and C5′(H)(n + 1)⋮O2′(n). These interactions may affect both backbone rigidity and thermodynamic stability of RNA helical structures. © 1999 John Wiley & Sons, Inc. Biopoly 50: 287–302, 1999     

The flanking sequence contributes to the immobilisation of spermine at the G-quadruplex in the NHE (nuclease hypersensitivity element) III1 of the c-Myc promoter

Defining the molecular basis of the DNA sequence selectivity of polyamine binding is central to understanding polyamine-dependent gene expression. We have studied, by selective NMR experiments, the variation of spermine mobility and conformation in the presence of G-quadruplexes formed by sequences of the purine-rich strand of the c-Myc promoter, nuclease hypersensitivity element III₁ (NHE III₁). All the NHE quadruplexes restrict spermine mobility and induce a spermine conformational change but the most effective immobilisation occurs when all five G-tracts of the NHE III₁ are present. This suggests structure within the nucleotides flanking the G-quadruplex has a role in immobilising spermine.     

Binding of spermine and ifenprodil to a purified,soluble regulatory domain of the N‐methyl‐d‐aspartate receptor

The binding of spermine and ifenprodil to the amino terminal regulatory (R) domain of the N‐methyl‐D ‐aspartate receptor was studied using purified regulatory domains of the NR1, NR2A and NR2B subunits, termed NR1‐R, NR2A‐R and NR2B‐R. The R domains were over‐expressed in Escherichia coli and purified to near homogeneity. The K_d values for binding of [¹⁴C]spermine to NR1‐R, NR2A‐R and NR2B‐R were 19, 140, and 33 μM, respectively. [³H]Ifenprodil bound to NR1‐R (K_d, 0.18 μM) and NR2B‐R (K_d, 0.21 μM), but not to NR2A‐R at the concentrations tested (0.1–0.8 μM). These K_d values were confirmed by circular dichroism measurements. The K_d values reflected their effective concentrations at intact NR1/NR2A and NR1/NR2B receptors. The results suggest that effects of spermine and ifenprodil on NMDA receptors occur through binding to the regulatory domains of the NR1, NR2A and NR2B subunits. The binding capacity of spermine or ifenprodil to a mixture of NR1‐R and NR2A‐R or NR1‐R and NR2B‐R was additive with that of each individual R domain. Binding of spermine to NR1‐R and NR2B‐R was not inhibited by ifenprodil and vice versa, indicating that the binding sites for spermine and ifenprodil on NR1‐R and NR2B‐R are distinct.     

Lipopolysaccharide‐bound structure of the antimicrobial peptide cecropin P1 determined by nuclear magnetic resonance spectroscopy

Antimicrobial peptides (AMPs) are components of the innate immune system and may be potential alternatives to conventional antibiotics because they exhibit broad‐spectrum antimicrobial activity. The AMP cecropin P1 (CP1), isolated from nematodes found in the stomachs of pigs, is known to exhibit antimicrobial activity against Gram‐negative bacteria. In this study, we investigated the interaction between CP1 and lipopolysaccharide (LPS), which is the main component of the outer membrane of Gram‐negative bacteria, using circular dichroism (CD) and nuclear magnetic resonance (NMR). CD results showed that CP1 formed an α‐helical structure in a solution containing LPS. For NMR experiments, we expressed ¹⁵N‐labeled and ¹³C‐labeled CP1 in bacterial cells and successfully assigned almost all backbone and side‐chain proton resonance peaks of CP1 in water for transferred nuclear Overhauser effect (Tr‐NOE) experiments in LPS. We performed ¹⁵N‐edited and ¹³C‐edited Tr‐NOE spectroscopy for CP1 bound to LPS. Tr‐NOE peaks were observed at the only C‐terminal region of CP1 in LPS. The results of structure calculation indicated that the C‐terminal region (Lys15–Gly29) formed the well‐defined α‐helical structure in LPS. Finally, the docking study revealed that Lys15/Lys16 interacted with phosphate at glucosamine I via an electrostatic interaction and that Ile22/Ile26 was in close proximity with the acyl chain of lipid A. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.     

An investigation of the dynamics of spermine bound to duplex and quadruplex DNA by <Superscript>13</Superscript>C NMR spectroscopy

A detailed analysis of the ¹³C relaxation of ¹³C-labelled spermine bound to duplex and quadruplex DNA is presented. T₁, T₂ and heteronuclear NOE data were collected at four ¹³C frequencies (75.4, 125.7, 150.9 and 201.2 MHz). The data were analyzed in terms of a frequency-dependent order parameter, S ²(ω), to estimate the generalized order parameter and the contributions to the relaxation from different motional frequencies in the picosecond–nanosecond timescale and from any exchange processes that may be occurring on the microsecond–millisecond timescale. The relaxation data was surprisingly similar for spermine bound to two different duplexes and a linear parallel quadruplex. Analysis of the relaxation data from these complexes confirmed the conclusions of previous studies that the dominant motion of spermine is independent of the macroscopic tumbling of the DNA and has an effective correlation time of ∼50 ps. In contrast, spermine bound to a folded antiparallel quadruplex had faster relaxation rates, especially R ₂. As with the other complexes, a fast internal motion of the order of 50 ps makes a substantial contribution to the relaxation. The generalized order parameter for spermine bound to duplex DNA and the linear quadruplex is small but is larger for spermine bound to the folded quadruplex. In the latter case, there is evidence for exchange between at least two populations of spermine occurring on the microsecond–millisecond timescale. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Association of putrescine,spermidine, spermine,and GABA with structural elements of brain cells

The present experiments are the first survey of the association of endogenous and exogenous putrescine, spermidine, and spermine with subcellular structures of rat brain cortex. The differences of distribution in subfractions obtained from salt-free and salt-containing density gradients were studied, with the following results: (1) In contrast with liver preparation, putrescine and the polyamines spermidine and spermine are not distributed in parallel with RNA. (2) In salt-containing media, putrescine and the polyamines were preferentially associated with synaptosomes and with synaptosomal membranes. Significant association with myelin constituents was observed only in salt-free media. (3) Exogenous putrescine and the polyamines were less firmly attached to synaptosomes and to synaptosomal membrane fractions than the endogenous amines. There is good evidence for similar subcellular localizations of putrescine and GABA. Putrescine seems to be entrapped in the nerve endings. (4) Uptake studies with crude mitochondria under conditions of high-affinity uptake showed no temperature-sensitive component of polyamine accumulation in synaptosomes, in contrast with GABA, monoacetylputrescine, and ornithine. (5) Polyamines bound to myelin constituents or mitochondria could be displaced by a 200-fold concentration of nonradioactive amines; this was not the case with polyamines bound to synaptosomes. Mg²⁺ did not effectively compete with spermine for binding sites at synaptic regions. (6) Electrical stimulation and stimulation by mono- and bivalent cations did not change the concentrations of the polyamines and GABA in guinea pig cortex. (7) There is no evidence for a neurotransmitter role of putrescine, spermidine, or spermine, although these compounds might function as modulators of neurotransmission.     

Inhibition of acetylpolyamine and spermine oxidases by the polyamine analogue chlorhexidine

Acetylpolyamine and spermine oxidases are involved in the catabolism of polyamines. The discovery of selective inhibitors of these enzymes represents an important tool for the development of novel anti-neoplastic drugs. Here, a comparative study on acetylpolyamine and spermine oxidases inhibition by the polyamine analogue chlorhexidine is reported. Chlorhexidine is an antiseptic diamide, commonly used as a bactericidal and bacteriostatic agent. Docking simulations indicate that chlorhexidine binding to these enzymes is compatible with the stereochemical properties of both acetylpolyamine oxidase and spermine oxidase active sites. In fact, chlorhexidine is predicted to establish several polar and hydrophobic interactions with the active site residues of both enzymes, with binding energy values ranging from ?7.6 to ?10.6 kcal/mol. In agreement with this hypothesis, inhibition studies indicate that chlorhexidine behaves as a strong competitive inhibitor of both enzymes, values of K_i being 0.10 μM and 0.55 μM for acetylpolyamine oxidase and spermine oxidase, respectively.     

Interactions of polyamines with the DNA octamers d(m5CG)4 and d(GGAATTCC): A 1H‐NMR investigation

The interaction of two DNA octamers, d(m⁵CG)₄ and d(GGAATTCC), with the polyamines spermine⁴⁺ and spermidine³⁺, has been studied by means of ¹H‐nmr nuclear Overhauser effect (NOE) difference measurements. The experiments were performed at 10°C and for a polyamine charge to DNA charge (i.e., phosphate) ratio of 0.4, where the solution of d(m⁵CG)₄ contains about 50% Z‐form of the DNA. The results show that the polyamine intramolecular NOEs for the protons on the propyl chains are similarly negative with the two oligonucleotides, while those on the butyl chain show slightly more negative NOE with d(m⁵CG)₄ than with d(GGAATTCC). The fully N‐methylated analogues of spermine (Me₁₀Spn⁴⁺) and spermidine (Me₈Spd³⁺) as well as the diamines 1,3‐diaminopropane (DAP²⁺) and 1,4‐diaminobutane (putrescine²⁺) have been studied for the ability to transform d(m⁵CG)₄ from the B‐ to the Z‐form. ¹H‐nmr spectra showed the order spermine⁴⁺ > spermidine³⁺ > Me₁₀Spn⁴⁺ > Me₈Spd³⁺ > 1,3‐diaminopropane²⁺ > putrescine²⁺, with spermine showing the largest relative amount of Z‐DNA. ¹H‐nmr pulsed‐gradient self‐diffusion measurements of the triamines showed a large difference in the interaction of Spd and Me₈Spd with the two different duplexes. With the same duplex (either of the two), however, no difference between Spd and Me₈Spd can be seen. Within a two‐state model this is interpreted as a larger fraction of bound polyamines with d(m⁵CG)₄ than with d(GGAATTCC). © 1999 John Wiley & Sons, Inc. Biopoly 49: 41–53, 1999     

Inhibition of the polyadenylation reaction in vitro by polyamines

The effect of polyamines on the polyadenylation reaction in vitro was investigated. Varying concentrations of spermine were added to the reaction catalyzed by purified poly(A) polymerase using rat liver nuclear RNA, poly(A), Escherichia coli tRNA or (Ap)₃A as exogenous primers. The enzyme activity decreased progressively with increasing concentrations of polyamines; complete inhibition was obtained at 0.4 and 1.2 mm spermine for the nuclear RNA- and poly(A)-primed reactions, respectively. No inhibition was observed for the (Ap)₃A-primed reaction. Spermidine and putrescine also inhibited polyadenylation but to a lesser extent than spermine. The degree of inhibition by spermine was related to the polynucleotide primer concentrations. Spermine prevented polyadenylation by binding to the primer but not to the poly(A) polymerase molecule as shown by the migration of [¹⁴C]spermine through glycerol gradients after preincubation with enzyme or tRNA. At concentrations inhibitory to polyadenylation in vitro, spermine could stimulate the DNA-dependent RNA synthesis catalyzed by RNA polymerase II. The present study suggests that low levels of polyamines could be used as specific inhibitors of the poly(A) synthesis in vitro.     

Occurrence of spermine in chromatin of Zea mays

Chromatin prepared from maize shoot tips using as extraction medium including quinacrine as an inhibitor of polyamine oxidase, contained 1.6 pmol spermidine g DNA^-1 and 14.8 pmol spermine g DNA^-1, respectively. This represented 0.1% spermidine and 3.7% spermine as compared with the content of those amines in the whole tissue. No putrescine was detectable in the chromatin preparation. When contamination of polyamines in the preparation was determined by the addition of labeled polyamines to the extraction medium, the ratio of the polyamines in the preparation to those in the extraction medium was 0.1% spermidine and 0.7% spermine, respectively. Spermine in the chromatin preparation was almost fully solubilized by a DNase-treatment, but spermidine was less easily solubilized. Most of the spermine associated with the chromation is chromatin-specific.     

Tpo1‐mediated spermine and spermidine export controls cell cycle delay and times antioxidant protein expression during the oxidative stress response

Cells counteract oxidative stress by altering metabolism, cell cycle and gene expression. However, the mechanisms that coordinate these adaptations are only marginally understood. Here we provide evidence that timing of these responses in yeast requires export of the polyamines spermidine and spermine. We show that during hydrogen peroxide (H₂O₂) exposure, the polyamine transporter Tpo1 controls spermidine and spermine concentrations and mediates induction of antioxidant proteins, including Hsp70, Hsp90, Hsp104 and Sod1. Moreover, Tpo1 determines a cell cycle delay during adaptation to increased oxidant levels, and affects H₂O₂ tolerance. Thus, central components of the stress response are timed through Tpo1‐controlled polyamine export.     

High-Field NMR and Circular Dichroism Solvent-Dependent Conformational Studies of the Bradykinin C-Terminal Tetrapeptide Ser-Pro-Phe-Arg

Abstract

The conformational properties of the tetrapeptide Ser¹-Pro²-Phe³-Arg⁴, the C-terminal fragment of the nonapeptide hormone bradykinin, have been studied by circular dichroism and two-dimensional NMR techniques. Measurements of coupling constants, NH temperature dependence rates and nuclear Overhauser effects (performed with rotating frame nuclear Overhauser spectroscopy, ROESY) in H₂O and CD₃OH/D₂O (80/20, v/v) reveal different conformations in the corresponding solvent. In aqueous solution the molecule exists in a random conformation or as an average of several conformations in rapid exchange. In CD₃OH/D₂O, however, the conformation is well-defined. The backbone of the peptide is extended, and the side-chains of Phe³ and Arg⁴ exhibit unusual rigidity for a peptide of this size. Evidently, the secondary structure is stabilized by a charge interaction between the guanidino group of Arg⁴ and the terminal carboxyl group, since experiments at various pH's show clearly that the definition of conformation decreases strongly upon protonation of the carboxyl function. A NH₃ ⁺(Ser¹)-COO^?(Arg⁴) salt bridge, as well as any form of turn stabilized by hydrogen bonds can be ruled out with certainty.     

Nuclear magnetic resonance studies on oligo-deoxyribonucleotides containing thedam methylation site GATC. Adenine methylation of d(GGATCC) does not change the helix geometry

We have studied the conformation of two hexanucleotides d(GGATCC) and d(GGm⁶ATCC) using proton nuclear magnetic resonance. Nuclear Overhauser effect measurements show that d(GGATCC) assumes a normal right handed B helix. The single and double strand resonances are in fast exchange on a proton nuclear magnetic resonance time scale. For d(GGm⁶ATCC), up to the Tm separate resonances are observed for each state, indicating slow exchange, though above the Tm it becomes more rapid. The orientation of the adenosine methyl-amino group, preferentiallycis to N¹, hinders base pair formation.The connectivities of the resonances of the two states were established by saturation transfer experiments. At 0°C irradiation of the m⁶ A-T imino proton gives an nuclear Overhauser effect to AH² showing that base pairing is Watson-Crick. Intra and interresidue nuclear Overhauser effects starting from the 3′ terminus show that the helix is right handed and in the B-form.The results on the two oligomers demonstrate that adenosine methylation induces little or no change in the conformation of the helix, but reduces the Tm from 45° to 32°C and slows the opening and closing of the m₆A.T base pair by a factor of about 100.     

Formation of β-alanine from spermine and spermidine in maize shoots

Spermine, spermidine or 1,3-diaminopropane fed to Zea mays shoots caused an accumulation of β-alanine. Furthermore, spermine 4 HCl [3-aminopropyl-3-³H(C)] was metabolized in the shoots to β-alanine. These results are consistent with the formation of β-alanine from polyamines by oxidative degradation.     

The identification of spermine binding sites in 16S rRNA allows interpretation of the spermine effect on ribosomal 30S subunit functions

A photoreactive analogue of spermine, N¹-azidobenzamidino (ABA)-spermine, was covalently attached after irradiation to Escherichia coli 30S ribosomal subunits or naked 16S rRNA. By means of RNase H digestion and primer extension, the cross-linking sites of ABA-spermine in naked 16S rRNA were characterised and compared with those identified in 30S subunits. The 5′ domain, the internal and terminal loops of helix H24, as well as the upper part of helix H44 in naked 16S rRNA, were found to be preferable binding sites for polyamines. Association of 16S rRNA with ribosomal proteins facilitated its interaction with photoprobe, except for 530 stem–loop nt, whose modification by ABA-spermine was abolished. Association of 30S with 50S subunits, poly(U) and AcPhe-tRNA (complex C) further altered the susceptibility of ABA-spermine cross-linking to 16S rRNA. Complex C, modified in its 30S subunit by ABA-spermine, reacted with puromycin similarly to non-photolabelled complex. On the contrary, poly(U)-programmed 70S ribosomes reconstituted from photolabelled 30S subunits and untreated 50S subunits bound AcPhe-tRNA more efficiently than untreated ribosomes, but were less able to recognise and reject near cognate aminoacyl-tRNA. The above can be interpreted in terms of conformational changes in 16S rRNA, induced by the incorporation of ABA-spermine.     

Evaluation of Staphylococcus aureus DNA aptamer by enzyme‐linked aptamer assay and isothermal titration calorimetry

To monitor the specificity of Staphylococcus aureus aptamer (SA‐31) against its target cell, we used enzyme‐linked aptamer assay. In the presence of target cell, horseradish peroxidase–conjugated streptavidin bound to biotin‐labeled SA‐31 showed specific binding to S   aureus among 3 different bacteria with limit of detection of 10³ colony‐forming unit per milliliter. The apparent K _a was 1.39 μM⁻¹ ± 0.3 μM⁻¹. The binding of SA‐31 to membrane proteins extracted from cell surface was characterized using isothermal titration calorimetry, and the effect of changes in binding temperature and salt concentrations of binding buffer was evaluated based on thermodynamic parameters (K _a, ΔH , and ΔG ). Since binding of aptamer to its targets solely depends on its 3‐dimensional structure under experimental conditions used in selection process, the change in temperature and ion concentration changed the affinity of SA‐31 to its target on surface of bacteria. At 4°C, SA‐31 did not show an affinity to its target with poor heat change upon injection of membrane fraction to aptamer solution. However, the apparent association constants of SA‐31 slightly varied from K _a = 1.56 μM⁻¹ ± 0.69 μM⁻¹ at 25°C to K _a = 1.03 μM⁻¹ ± 0.9 μM⁻¹ at 37°C. At spontaneously occurring exothermic binding reactions, affinities of S  aureus aptamer to its target were also 9.44 μM⁻¹ ± 0.38 μM⁻¹ at 50mM, 1.60 μM⁻¹ ± 0.11 μM⁻¹ at 137mM, and 3.28 μM⁻¹ ± 0.46 μM⁻¹ at 200 mM of salt concentration. In this study, it was demonstrated that enzyme‐linked aptamer assay and isothermal titration calorimetry were useful tools for studying the fundamental binding mechanism between a DNA aptamer and its target on the outer surface of S  aureus .     

Molecular dynamics simulation of the induced‐fit binding process of DNA aptamer and L‐argininamide

Aptamers are rare functional nucleic acids with binding affinity to and specificity for target ligands. Recent experiments have lead to the proposal of an induced‐fit binding mechanism for L ‐argininamide (Arm) and its binding aptamer. However, at the molecular level, this mechanism between the aptamer and its coupled ligand is still poorly understood. The present study used explicit solvent molecular dynamics (MD) simulations to examine the critical bases involved in aptamer‐Arm binding and the induced‐fit binding process at atomic resolution. The simulation results revealed that the Watson‐Crick pair (G10‐C16), C9, A12, and C17 bases play important roles in aptamer‐Arm binding, and that binding of Arm results in an aptamer conformation optimized through a general induced‐fit process. In an aqueous solution, the mechanism has the following characteristic stages: (a) adsorption stage, the Arm anchors to the binding site of aptamer with strong electrostatic interaction; (b) binding stage, the Arm fits into the binding site of aptamer by hydrogen‐bond formation; and (c) complex stabilization stage, the hydrogen bonding and electrostatic interactions cooperatively stabilize the complex structure. This study provides dynamics information on the aptamer‐ligand induced‐fit binding mechanism. The critical bases in aptamer‐ligand binding may provide a guideline in aptamer design for molecular recognition engineering.