RNA-Binding Potential of Protoberberine Alkaloids: Spectroscopic and Calorimetric Studies on the Binding of Berberine, Palmatine, and Coralyne to Protonated RNA Structures

β-Carboline alkaloids are present in medicinal plants such as Peganum harmala L., which have been used as folk medicine in anticancer therapy. Recently, they have drawn attention because of their antitumor activities. Despite considerable interest and investigations on alkaloid-DNA complexes, reports on alkaloid-RNA interaction are very limited. This study is the first attempt to investigate the binding of β-carboline alkaloids (harmine, harmane, harmaline, harmalol, and tryptoline) with yeast RNA. The effect of alkaloid complexation on RNA aggregation and condensation was investigated in aqueous solution at physiological conditions, using constant RNA concentration (6.25 mM) and various alkaloid:polynucleotide (phosphate) ratios of 1:240, 1:160, 1:80, 1:40, 1:20, 1:10, 1:5, 1:2, and 1:1. Fourier transform infrared and UV-visible spectroscopic methods were used to determine the ligand-binding modes, the binding constants, and the stability of alkaloid-RNA complexes in aqueous solution. Spectroscopic evidence showed major binding of alkaloids to RNA with overall binding constants of K(harmine)-RNA = 2.95 × 10⁷ M⁻¹, K(harmane)-RNA = 5.62 × 10⁵ M⁻¹, K(harmaline)-RNA = 7.47 × 10⁵ M⁻¹, K(harmalol)-RNA = 4.32 × 10⁵ M⁻¹, and K(tryptoline)-RNA = 3.21 × 10⁵ M⁻¹. The affinity of alkaloids-RNA binding is in the order of harmine > harmaline > harmane > harmalol > tryptoline. No biopolymer secondary structural changes were observed upon alkaloid interaction and RNA remains in the A-family structure in these complexes.

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“…Similar binding constants were estimated for other alkaloids-RNA complexes (Maidul Islam and Kumar, 2009). …”

Section: Stability Of Alkaloid-rna Complexessupporting

confidence: 75%

Interaction of β-Carboline Alkaloids with RNA

Nafisi

Malekabady

Khalilzadeh

2010

DNA and Cell Biology

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“…To elucidate this problem further, the binding reactions between RNA and flavonoids in an aqueous phase were studied using ITC. ITC is an important tool for measuring the thermodynamics of interactions between small molecules and biopolymers, in which all binding parameters (n, K, ΔH and ΔS) are simultaneously determined in a single ITC experiment, thus obtaining information that cannot be obtained by other methods [35]–[37]. In our research, analysis of the ITC measurements revealed two interactions to be primarily enthalpy driven.…”

Section: Discussionmentioning

confidence: 68%

Anti- Japanese-Encephalitis-Viral Effects of Kaempferol and Daidzin and Their RNA-Binding Characteristics

Zhang

Wú

et al. 2012

PLoS ONE

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BackgroundNew therapeutic tools and molecular targets are needed for treatment of Japanese encephalitis virus (JEV) infections. JEV requires an α-1 translational frameshift to synthesize the NS1' protein required for viral neuroinvasiveness. Several flavonoids have been shown to possess antiviral activity in vitro against a wide spectrum of viruses. To date, the antiviral activities of flavonol kaempferol (Kae) and isoflavonoid daidzin (Dai) against JEV have not been described.Methodology/Principal FindingsThe 50% cytotoxic concentration (CC50) and 50% effective concentration (EC50) against JEV were investigated in BHK21 cells by MTS reduction. Activity against viral genomic RNA and proteins was measured by real-time RT-PCR and western blotting. The frameshift site RNA-binding characterization was also determined by electrospray ionization mass spectrometry, isothermal titration calorimetry and autodocking analysis. EC50 values of Kae and Dai were 12.6 and 25.9 µM against JEV in cells pretreated before infection, whereas in cells infected before treatment, EC50 was 21.5 and 40.4 µM, respectively. Kae exhibited more potent activity against JEV and RNA binding in cells following internalization through direct inhibition of viral replication and protein expression, indicating that its antiviral activity was principally due to direct virucidal effects. The JEV frameshift site RNA (fsRNA) was selected as a target for assaying Kae and Dai. ITC of fsRNA revealed an apparent Kb value for Kae that was nine fold stronger than that for Dai. This binding was confirmed and localized to the RNA using ESI-MS and autodock analysis. Kae could form non-covalent complexes with fsRNA more easily than Dai could.Conclusions/SignificanceKae demonstrates more potent antiviral activity against JEV than does Dai. The mode of action of Kae as an anti-JEV agent seems to be related to its ability to inactivate virus by binding with JEV fsRNA.

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“…Understanding the mechanism(s) of action of berberine is of significant interest and requires further studies. Berberine is a known DNA and RNA binder [45], [11], [46], [47], and recently it was reported to target the bacterial division protein FtsZ [2]. The evidence supporting these findings was obtained mainly through biochemical and biophysical studies.…”

Section: Discussionmentioning

confidence: 96%

Genetic Evidence for Inhibition of Bacterial Division Protein FtsZ by Berberine

2010

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BackgroundBerberine is a plant alkaloid that is widely used as an anti-infective in traditional medicine. Escherichia coli exposed to berberine form filaments, suggesting an antibacterial mechanism that involves inhibition of cell division. Berberine is a DNA ligand and may induce filamentation through induction of the SOS response. Also, there is biochemical evidence for berberine inhibition of the cell division protein FtsZ. Here we aimed to assess possible berberine mechanism(s) of action in growing bacteria using genetics tools.Methodology/Principal FindingsFirst, we tested whether berberine inhibits bacterial growth through DNA damage and induction of the SOS response. The SOS response induced by berberine was much lower compared to that induced by mitomycin C in an SOS response reporter strain. Also, cell filamentation was observed in an SOS-negative E. coli strain. To test whether berberine inhibits FtsZ, we assessed its effects on formation of the cell division Z-rings, and observed a dramatic reduction in Z-rings in the presence of berberine. We next used two different strategies for RNA silencing of ftsZ and both resulted in sensitisation of bacteria to berberine, visible as a drop in the Minimum Inhibitory Concentration (MIC). Furthermore, Fractional Inhibitory Concentration Indices (FICIs) showed a high level of synergy between ftsZ silencing and berberine treatment (FICI values of 0.23 and 0.25 for peptide nucleic acid- and expressed antisense RNA-based silencing of ftsZ, respectively). Finally, over-expression of ftsZ led to a mild rescue effect in berberine-treated cells.ConclusionsThe results argue against DNA binding as the primary mechanism of action of berberine and support the hypothesis that its antibacterial properties are due to inhibition of the cell division protein FtsZ. In addition, the genetic approach used here provides a means to rapidly test the activity of other putative FtsZ inhibitors.

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RNA-Binding Potential of Protoberberine Alkaloids: Spectroscopic and Calorimetric Studies on the Binding of Berberine, Palmatine, and Coralyne to Protonated RNA Structures

Cited by 18 publications

References 50 publications

Interaction of β-Carboline Alkaloids with RNA

Interaction of β-Carboline Alkaloids with RNA

Anti- Japanese-Encephalitis-Viral Effects of Kaempferol and Daidzin and Their RNA-Binding Characteristics

Genetic Evidence for Inhibition of Bacterial Division Protein FtsZ by Berberine

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