Article pubs.acs.org/JPCB
Molecular Recognition of tRNA with 1‑Naphthyl Acetyl Spermine, Spermine, and Spermidine: A Thermodynamic, Biophysical, and Molecular Docking Investigative Approach Ayesha Kabir,† Devawati Dutta,‡ Chhabinath Mandal,§ and Gopinatha Suresh Kumar*,† Biophysical Chemistry Laboratory, Organic and Medicinal Chemistry Division, and ‡Cancer Biology and Inflammatory Disorder Division, CSIR-Indian Institute of Chemical Biology, 4, Raja S. C. Mullick Road, Kolkata 700032, India § National Institute of Pharmaceutical and Educational Research, Kolkata 700032, India Downloaded via TULANE UNIV on January 23, 2019 at 12:54:46 (UTC). See https://pubs.acs.org/sharingguidelines for options on how to legitimately share published articles.
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ABSTRACT: The role of tRNA in protein translational machinery and the influence of polyamines on the interaction of acylated and deacylated tRNA with ribosomes make polyamine−tRNA interaction conspicuous. We studied the interaction of two biogenic polyamines, spermine (SPM) and spermidine (SPD), with tRNAPhe and compared the results to those of the analogue 1-naphthyl acetyl spermine (NASPM). The binding affinity of SPM was comparable to that of NASPM; both were higher than that of SPD. The interactions led to significant thermal stabilization of tRNAPhe and an increase in the enthalpy of transition. All the interactions were exothermic in nature and displayed prominent enthalpy− entropy compensation behavior. The entropy-driven nature of the interaction, the structural perturbations observed, and docking results proved that the polyamines were bound in the groove of the anticodon arm of tRNAPhe. The amine groups of polyamines were involved in extensive electrostatic, H-bonding, and van der Waals interactions with tRNAPhe. The naphthyl group of NASPM showed an additional stacking interaction with G24 and G26 of tRNAPhe, which was absent in others. The results demonstrate that 1-naphthyl acetyl spermine can target the same binding sites as the biogenic polyamines without substituting for the functions played by them, which may lead to exhibition of selective anticancer cytotoxicity.
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INTRODUCTION RNA plays a central role in transcription, translation, protein functions, enzymatic processes, gene regulation, and also RNA interference.1−6 Therefore, RNA-binding small molecules may serve as leads for RNA-based antiviral and anticancer agents.7−11 Aminoglycoside antibiotics are one group of RNA-binding molecules that interact at the functional centers of the 16S rRNA.12−14 Transfer RNAs (tRNA) are one of the smallest RNA molecules; they function as cardinal decoding adaptors in the synthesis of proteins.15,16 tRNA has a wellcharacterized structure with a sequence of about 74−95 bases. The cloverleaf secondary structure of tRNA consists of four constant arms and an extra arm in the longer tRNAs. The molecule folds back on itself forming an L-shaped tertiary structure consisting of two segments of double helix.17 Even though tRNAs are mainly known for their central role in translation, they also engage in a number of nontranslational activities in bacteria, eukaryotes, and archaea.17,18 They play pivotal roles in stress signaling, adaptive translation, and complex human diseases.18 Additionally, the tRNA structure represents a unique site for recognition of small molecules, and such binding interactions have been documented through extensive studies.19−22 © 2016 American Chemical Society
Polyamines, on the other hand, regulate gene expression and cellular growth and activities through its interaction with nucleic acids and proteins.23−26 Cancerous cells often display formidable concentration of polyamines, whereas a waning in their concentration may lead to cellular growth suppression followed by apoptosis.27−31 For these reasons, understanding polyamine action and metabolism has been crucial for antiproliferative and anticancer drug development.32 Minor structural changes in the biogenic polyamines have yielded analogues with paramount biological relevance in recent times.32−35 The selective polyamine transporter of the cell may be utilized by a potent polyamine analogue to enter the cell and tussle with the targets of the natural polyamines.32−35 Considering the fact that the analogue does not replace the biogenic polyamines in proliferative function, it may display selective anticancer cytotoxicity.32−35 Bis(ethyl) polyamines are the most notable analogues that have ever been synthesized.34,35 Various reports have shown the potency of a number of other polyamine analogues against cancerous growth in Received: May 28, 2016 Revised: October 1, 2016 Published: October 3, 2016 10871
DOI: 10.1021/acs.jpcb.6b05391 J. Phys. Chem. B 2016, 120, 10871−10884
Article
The Journal of Physical Chemistry B
Figure 1. (A) Secondary and tertiary structures of tRNA, (B) chemical structure of spermine (SPM), 1-naphthyl acetyl spermine (NASPM), and spermidine (SPD).
numerous cell lines.34−40 Reports have shown that polyamines and their analogues interact with and stabilize tRNA.41−46 The interaction of natural polyamines with tRNA is pertinent for optimal translational preciseness and efficiency.46 Polyamines have been reported to attune protein synthesis and stimulate the binding interactions of acylated and deacylated tRNA to ribosomes.47,48 Thus, the vital and expansive nature of tRNAs make them crucial targets of drugs. For these reasons, it will be interesting to validate the interaction of polyamines and their analogue with tRNA. Our work on the comparative binding of biogenic polyamines and the SPM analogue, 1-naphthyl acetyl spermine (NASPM), with RNA and DNA polynucleotides has given us interesting results, where we observed different specificity behavior of the analogue NASPM over natural polyamines.49,50 The aim of this work is to widen our knowledge related to the comparative binding of polyamines and analogues with tRNAPhe. The tRNAPhe binding potency of NASPM is not yet known. NASPM (Figure 1) is a synthetic analogue of the Joro spider toxin, which has been reported to display long-lasting anticonvulsant effect on previously kindled rats.51,52 Thus, biophysical and calorimetric studies were performed to elucidate the structural, thermodynamic, and conformational aspects of the binding interaction. Additionally, molecular docking studies were carried out to evaluate the strength and mode of the interactions involved in binding.
of protein contamination, as indicated from the absorbance ratio at 260/280 nm. Each day, the polyamines were weighed and dissolved to obtain the polyamine solutions. The experiments were done in 20 mM [Na+] in citrate-phosphate buffer, pH 7.0, containing 10 mM Na2HPO4 and the desired pH was attained by using citric acid; 0.22 μM millipore filters (Millipore, India Pvt. Ltd, Bangalore, India) were used to filter the buffer solutions. Different amounts of NaCl were added to the buffer solution to obtain buffers of different salt concentrations required for the salt-dependent studies. Experimental Methods. Differential scanning calorimetry (DSC) is a dependable methodology used to measure the heat energy uptake of a sample during controlled increase (or decrease) in temperature. The measurements were performed with a Microcal VP DSC unit (MicroCal, LLC., Northampton, MA, now Malvern Instruments Ltd., Malvern, U.K.).53 In a set of continuous DSC scans, the cells were loaded with the buffer and equilibrated at 25 °C for 15 min. It was then scanned from 25 to 120 °C, at approximately 30 psi pressure, at a scan rate of 60 °C/h. The buffer scans were repeated till the baseline was reproducible (noise specification
