Gopinatha Suresh Kumar

Therapeutic potential of nucleic acid-binding isoquinoline alkaloids: Binding aspects and implications for drug design

Isoquinoline alkaloids represent a group of natural products with remarkable importance in the contemporary biomedical research and drug discovery programs. Several members of this group exhibit immense pharmacological and biological properties, including potential anticancer properties. Although the molecular targets of these alkaloids are not yet clearly delineated, extensive research in this area continues to build up new data that are clinically exploitable. The gross structural features of many of the members DNA interaction are more or less clear, but the mystery still remains on many aspects of their binding, including specificity and energetics. RNA‐binding aspects of these alkaloids are being elucidated. More recent advancements in analytical instrumentation have enabled clearer elucidation and correlation of the structural and energetic aspects of the interaction. In this review, we report up‐to‐date details of the interaction of berberine, palmatine, and jatrorrhizine of the protoberberine group, sanguinarine from the benzophananthridine group, and several of their synthetic derivatives, such as coralyne, berberrubine, palmatrubine, and jatrorubin with nucleic acids have been reviewed. These studies, taken together up to now, have led to interesting knowledge on the mode, mechanism, specificity of binding, and correlation between structural aspects and energetics enabling a complete set of guidelines for design of new drugs. In contemporary research, several derivatives of these natural alkaloids are being prepared and investigated in several laboratories for ultimate discovery of new compounds that can be used as effective therapeutic agents.

Spectroscopic and calorimetric studies on the binding of alkaloids berberine, palmatine and coralyne to double stranded RNA polynucleotides.

The interaction of two natural protoberberine plant alkaloids berberine and palmatine and a synthetic derivative coralyne to three double stranded ribonucleic acids, poly(A). poly(U), poly(I).poly(C) and poly(C).poly(G) was studied using various biophysical techniques. Absorbance and fluorescence studies showed that the alkaloids bound cooperatively to these RNAs with the binding affinities of the order 10(4) M(-1). Circular dichroic results suggested that the conformation of poly(A). poly(U) was perturbed by all the three alkaloids, that of poly(I).poly(C) by coralyne only and that of poly(C).poly(G) by none. Fluorescence quenching studies gave evidence for partial intercalation of berberine and palmatine and complete intercalation of coralyne to these RNA duplexes. Isothermal titration calorimetric studies revealed that the binding was characterized by negative enthalpy and positive entropy changes and the affinity constants derived were in agreement with the overall binding affinity from spectral data. The binding of all the three alkaloids considerably stabilized the melting of poly(A). poly(U) and poly(I).poly(C) and the binding data evaluated from the melting data were in agreement with that obtained from other techniques. The overall binding affinity of the alkaloids to these double stranded RNAs varied in the order, berberine = palmatine < coralyne. The temperature dependence of the enthalpy changes afforded large negative values of heat capacity changes for the binding of palmatine and coralyne to poly(A).poly(U) and of coralyne to poly(I).poly(C), suggesting substantial hydrophobic contribution in the binding process. Further, enthalpy-entropy compensation was also seen in almost all the systems that showed binding. These results further advance our understanding on the binding of small molecules that are specific binders to double stranded RNA sequences.

Interaction of berberine, palmatine, coralyne, and sanguinarine to quadruplex DNA: a comparative spectroscopic and calorimetric study.

BACKGROUND Interaction of isoquinoline alkaloids berberine, palmatine, coralyne and sanguinarine with human telomeric quadruplex DNA, dAGGG(TTAGGG)(3), has been investigated and compared with ethidium. METHODS Biophysical techniques such as absorption, fluorescence, circular dichroism, optical melting and microcalorimetry have been used. RESULTS Absorption and fluorescence studies revealed noncooperative 1:1 binding for all the molecules. Coralyne showed highest affinity (10(6) M(-1)) and for others it was ~10(5) M(-1). The binding affinity varied as coralyne>sanguinarine>berberine>palmatine. Ethidium showed affinity close to sanguinarine. Comparative fluorescence quenching and polarization anisotropy of the emission spectra gave evidence for a stronger stacking interaction of coralyne and sanguinarine compared to berberine and palmatine. Circular dichroic spectral perturbations were similar in all the cases, but a strong induced circular dichroism for the bound molecules was observed only for coralyne and sanguinarine. The interaction of all the alkaloids was exothermic. Binding of coralyne and sanguinarine was predominantly enthalpy driven while that of berberine and palmatine was entropy driven. Heat capacity values of -169, -198, -105 and -95cal/molK, respectively, for coralyne, sanguinarine, berberine, and palmatine suggested significant differences in the hydrophobic contribution to the binding. CONCLUSIONS This study presents a complete structural and thermodynamic profile of the binding of isoquinoline alkaloids with G-quadruplex. GENERAL SIGNIFICANCE These results suggest strong and specific binding of these molecules to the G-quadruplex and highlight the differences in their structure in the interaction profile.

Self-structure induction in single stranded poly(A) by small molecules: Studies on DNA intercalators, partial intercalators and groove binding molecules

Self-structure induction in single stranded poly(A) has been one typical example of the various ways that could be used to modulate nucleic acid structural aspects through binding of small molecules. For the first time, the interaction between a series of small molecules and poly(A) has been investigated to understand the nature of the structural features in DNA binding small molecules that could be responsible for the formation of self-structure in single stranded poly(A) molecules. Classical intercalators like ethidium, coralyne, quinacrine and proflavine, partial intercalators like berberine and palmatine and classical minor groove binders like hoechst 33258 and DAPI have been chosen for this study. The binding of each of these molecules to poly(A) has been characterized by absorption spectral titration, job plot and isothermal titration calorimetry. Self-structure formation was monitored from circular dichroic melting, optical melting and differential scanning calorimetry. The results revealed that while all the intercalators studied induced self-structure formation, partial intercalators did not induce the same in poly(A). Of the two classical DNA minor groove binding molecules investigated, hoechst was effective in inducing self-structure while DAPI was ineffective. Self-structure induction in poly(A) was observed to be directly linked to the cooperative binding of the molecules to poly(A) in that all the molecules that bound cooperatively induced self-structure in poly(A). Structural and thermodynamic aspects of the interaction leading to self-structure formation are described.

Interaction of Isoquinoline Alkaloids with an RNA Triplex: Structural and Thermodynamic Studies of Berberine, Palmatine, and Coralyne Binding to Poly(U).Poly(A)*Poly(U)

The interaction of two natural protoberberine alkaloids berberine and palmatine and the synthetic derivative coralyne with the RNA triplex poly(U).poly(A)(*)poly(U) was studied using various biophysical and calorimetric techniques. All the three alkaloids bind noncooperatively to the triplex. The affinity of berberine and palmatine was in the order of 10(5) M(-1), while that of coralyne was one order higher as inferred from spectroscopic studies. The alkaloids stabilized the Hoogsteen base-paired third strand of the triplex without affecting the stability of the duplex. Fluorescence quenching and viscosity studies gave convincing evidence for the partial intercalation of berberine and palmatine and a true intercalative binding of coralyne to the triplex. This was further supported from the significant polarization of the emission spectra of the complex and the energy transfer from the base triplets to the alkaloids. Circular dichroic studies suggested that the conformation of the triplex was perturbed significantly by the binding of the alkaloids, being more by coralyne compared to berberine and palmatine and also evidenced by the generation of strong induced optical activity in the bound coralyne molecules. Isothermal titration calorimetric studies revealed that the binding to the triplex was favored by a predominantly large negative enthalpy change (DeltaH degrees = -5.42 kcal/mol) with small favorable entropy contribution (TDeltaS degrees = 2.02 kcal/mol) in berberine, favored by almost equal negative enthalpy (DeltaH degrees = -3.93 kcal/mol) and entropy changes (TDeltaS degrees = 3.89 kcal/mol) in palmatine and driven by predominant entropy contributions (DeltaH degrees = -1.84 and TDeltaS degrees = 7.44 kcal/mol) in coralyne. These results advance our knowledge on the binding of small molecule isoquinoline alkaloids that are specific binders of RNA structures, particularly triplexes.

Spectroscopic and Thermodynamic Studies on the Binding of Sanguinarine and Berberine to Triple and Double Helical DNA and RNA Structures

Abstract A comparative study on the interaction of sanguinarine and berberine with DNA and RNA triplexes and their parent duplexes was performed, by using a combination of spectrophotometric, UV thermal melting, circular dichroic and thermodynamic techniques. Formation of the DNA and RNA triplexes was confirmed from UV-melting and circular dichroic measurements. The interaction process was characterized by increase of thermal melting temperature, perturbation in circular dichroic spectrum and the typical hypochromic and bathochromic effects in the absorption spectrum. Scatchard analysis indicated that both the alkaloids bound to the triplex and duplex structures in a non-cooperative manner and the binding was stronger to triplexes than to parent duplexes. Thermal melting studies further indicated that sanguinarine stabilized the Hoogsteen base paired third strand of both DNA and RNA triplexes more tightly compared to their Watson-Crick strands, while berberine stabilized the third strand only without affecting the Watson-Crick strand. However, sanguinarine stabilized the parent duplexes while no stabilization was observed with berberine under identical conditions. Circular dichroic studies were also consistent with the observation that perturbations of DNA and RNA triplexes were more compared to their parent duplexes in presence of the alkaloids. Thermodynamic data revealed that binding of sanguinarine and berberine to triplexes (T·AxT and U·AxU) and duplexes (A·T and A·U) showed negative enthalpy changes and positive entropy changes but that of sanguinarine to C·GxC+ triplex and G·C duplex exhibited negative enthalpy and negative entropy changes. Taken together, these results suggest that both sanguinarine and berberine can bind and stabilize the DNA and RNA triplexes more strongly than their respective parent duplexes.

Berberine–DNA complexation: New insights into the cooperative binding and energetic aspects

The equilibrium binding of the cytotoxic plant alkaloid berberine to various DNAs and energetics of the interaction have been studied. At low ratios of bound alkaloid to base pair, the binding exhibited cooperativity to natural DNAs having almost equal proportions of AT and GC sequences. In contrast, the binding was non-cooperative to DNAs with predominantly high AT or GC sequences. Among the synthetic DNAs, cooperative binding was observed with poly(dA).poly(dT) and poly(dG).poly(dC) while non-cooperative binding was seen with poly(dA-dT).poly(dA-dT) and poly(dG-dC).poly(dG-dC). Both cooperative and non-cooperative bindings were remarkably dependent on the salt concentration of the media. Linear plots of ln K(a) versus [Na(+)] for poly(dA).poly(dT) and poly(dA-dT).poly(dA-dT) showed the release of 0.56 and 0.75 sodium ions respectively per bound alkaloid. Isothermal titration calorimetry results revealed the binding to be exothermic and favoured by both enthalpy and entropy changes in all DNAs except the two AT polymers and AT rich DNA, where the same was predominantly entropy driven. Heat capacity values (DeltaCp(o)) of berberine binding to poly(dA).poly(dT), poly(dA-dT).poly(dA-dT), Clostridium perfringens and calf thymus DNA were -98, -140, -120 and -110 cal/mol K respectively. This study presents new insights into the binding dependent base pair heterogeneity in DNA conformation and the first complete thermodynamic profile of berberine binding to DNAs.

Interaction of the anticancer plant alkaloid sanguinarine with bovine serum albumin.

Background Interaction of the iminium and alkanolamine forms of sanguinarine with bovine serum albumin (BSA) was characterized by spectroscopic and calorimetric techniques. Methodology/Principal Findings Formation of strong complexes of BSA with both iminium and alkanolamine forms was revealed from fluorescence quenching of sanguinarine. Binding parameters calculated from Stern-Volmer quenching method revealed that the neutral alkanolamine had higher affinity to BSA compared to the charged iminium form. Specific binding distances of 3.37 and 2.38 nm between Trp 212 (donor) and iminium and alkanolamine forms (acceptor), respectively, were obtained from Forster resonance energy transfer studies. Competitive binding using the site markers warfarin and ibuprofen, having definite binding sites, demonstrated that both forms of sanguinarine bind to site I (subdomain IIA) on BSA. Sanguinarine binding alters protein conformation by reducing the α-helical organization and increasing the coiled structure, indicating a small but definitive partial unfolding of the protein. Thermodynamic parameters evaluated from isothermal titration calorimetry suggested that the binding was enthalpy driven for the iminium form but favoured by negative enthalpy and strong favourable entropy contributions for the alkanolamine form, revealing the involvement of different molecular forces in the complexation. Conclusions/Significance The results suggest that the neutral alkanolamine form binds to the protein more favourably compared to the charged iminium, in stark contrast to the reported DNA binding preference of sanguinarine.

Polymorphic Nucleic Acid Binding of Bioactive Isoquinoline Alkaloids and Their Role in Cancer

Bioactive alkaloids occupy an important position in applied chemistry and play an indispensable role in medicinal chemistry. Amongst them, isoquinoline alkaloids like berberine, palmatine and coralyne of protoberberine group, sanguinarine of the benzophenanthridine group, and their derivatives represent an important class of molecules for their broad range of clinical and pharmacological utility. In view of their extensive occurrence in various plant species and significantly low toxicities, prospective development and use of these alkaloids as effective anticancer agents are matters of great current interest. This review has focused on the interaction of these alkaloids with polymorphic nucleic acid structures (B-form, A-form, Z-form, HL-form, triple helical form, quadruplex form) and their topoisomerase inhibitory activity reported by several research groups using various biophysical techniques like spectrophotometry, spectrofluorimetry, thermal melting, circular dichroism, NMR spectroscopy, electrospray ionization mass spectroscopy, viscosity, isothermal titration calorimetry, differential scanning calorimetry, molecular modeling studies, and so forth, to elucidate their mode and mechanism of action for structure-activity relationships. The DNA binding of the planar sanguinarine and coralyne are found to be stronger and thermodynamically more favoured compared to the buckled structure of berberine and palmatine and correlate well with the intercalative mechanism of sanguinarine and coralyne and the partial intercalation by berberine and palmatine. Nucleic acid binding properties are also interpreted in relation to their anticancer activity.

Isoquinoline Alkaloids and their Binding with Polyadenylic Acid: Potential Basis of Therapeutic Action

After fifty years of DNA targeting through intercalators and groove binders and related studies now the current focus is in RNA targeting. Polyadenylic acid [poly(A)] tail of mRNA has been recently established as a potential drug target due to its significant role in the initiation of translation, maturation and stability of mRNA as well as in the production of alternate proteins in eukaryotic cells. Isoquinoline group of alkaloids have their importance in contemporary biomedical research and drug discovery programme due to extensive pharmacological and biological activity. Very recently some small molecule alkaloids of the isoquinoline group have been found to bind poly(A) with remarkably high affinity leading to self structure formation. The alkaloids have a high binding affinity towards single stranded poly(A) whereas their binding with double stranded poly(A) is weak. Among the alkaloids discussed here, berberine and coralyne are found to be capable to induce self-structure in poly(A). All the binding phenomena are characterized by electrostatic interaction between RNA and the alkaloids and the mode of binding revealed as full or partial intercalation. This review focuses on the structural and biological significance of poly(A) and the recent developments in the use of plant alkaloids and their synthetic analogs to control the structure and function of this RNA for the development of new alkaloid based molecules specifically targeted to poly(A) structures.