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ABSTRACT: The nucleocapsid protein (N) and the phosphoprotein (P) of nonsegmented negative-strand (NNS) RNA viruses interact with each other to accomplish two crucial events necessary for the viral replication cycle. First, the P protein binds to the aggregation prone nascent N molecules maintaining them in a soluble monomeric (N(0)) form (N(0)-P complex). It is this form that is competent for specific encapsidation of the viral genome. Second, the P protein binds to oligomeric N in the nucleoprotein complex (N-RNA-P complex), and thereby facilitates the recruitment of the viral polymerase (L) onto its template. All previous attempts to study these complexes relied on co-expression of the two proteins in diverse systems. In this study, we have characterised these different modes of N-P interaction in detail and for the first time have been able to reconstitute these complexes individually in vitro in the chandipura virus (CHPV), a human pathogenic NNS RNA virus. Using a battery of truncated mutants of the N protein, we have been able to identify two mutually exclusive domains of N involved in differential interaction with the P protein. An unique N-terminal binding site, comprising of amino acids (aa) 1-180 form the N(0)-P interacting region, whereas, C-terminal residues spanning aa 320-390 is instrumental in N-RNA-P interactions. Significantly, the ex-vivo data also supports these observations. Based on these results, we suggest that the P protein acts as N-specific chaperone and thereby partially masking the N-N self-association region, which leads to the specific recognition of viral genome RNA by N(0).
PLoS ONE 01/2012; 7(4):e34623. · 4.09 Impact Factor
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ABSTRACT: Chandipura virus, a member of the vesiculovirus genera, has been recently recognized as an emerging human pathogen. Previously, we have shown that Chandipura virus Nucleocapsid protein N is capable of binding to both specific viral leader RNA as well as non-viral RNA sequences, albeit in distinct monomeric and oligomeric states, respectively. Here, we distinguish the regions of N involved in oligomerization and RNA binding using a panel of deletion mutants. We demonstrate that deletion in the N-terminal arm completely abrogates self-association of N protein. Monomer N specifically recognizes viral leader RNA using its C-terminal 102 residues, while oligomerization generates an additional RNA binding surface involving the N-terminal 320 amino acids of N overlapping with a protease resistant core that is capable of forming nucleocapsid like structure and also binding heterogeneous RNA sequences. Finally, we propose a model to explain the mechanism of genome encapsidation of this important human pathogen.
Virology 11/2010; 407(1):33-42. · 3.35 Impact Factor
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ABSTRACT: The chemical reaction of Pt(II)(L(1))Cl(2) [L(1) = N-4-tolylpyridine-2-aldimine] with a bidentate N,S-donor atom ligand, 2-methylthioaniline, (HL(2)) in alkaline methanolic medium yielded a mixed ligand donor-acceptor complex, [Pt(II)(L(1))(L(2))]Cl, [1]Cl. The complex has been characterized by different spectroscopic and electrochemical techniques. The complex showed intense interligand charge transfer (ILCT) transition in the long wavelength region of UV-vis spectrum (>600 nm). The single-crystal X-ray structure of complex, [1]Cl·3.3H(2)O is reported. The cationic complex upon crystallization from aqueous methanol solvent produces an assembly of discrete, three dimensional (H(2)O)(20) guest moiety within the reference Pt-complex host lattice. The water assembly showed a unique type of aggregation of a distorted cube encapsulated by hydrogen bonded network of a twelve-water ring. The complex displayed one reversible cathodic response at -0.75 V and two irreversible anodic responses at 0.42 and 0.79 V versus Ag/AgCl reference electrode. The redox processes are characterized by EPR and spectroelectrochemistry. Density-functional theory calculations were employed to confirm the structural features and to support the spectral and redox properties of the complex. The square-planar complex has been found to intercalate DNA. Fluorescence spectroscopy, circular dichroism, cyclic voltammetry, viscosity measurements, together with DNA melting studies have been employed to characterize the binding of [1]Cl with calf thymus DNA. Agarose gel electrophoresis indicates that the complex cleaves supercoiled (SC) pUC19 plasmid DNA to its nicked circular (NC) form via singlet oxygen. As determined by a MTT assay, [1]Cl exhibits significant cytotoxicity with IC(50) value 58 μM.
Dalton Transactions 10/2010; 39(40):9514-22. · 3.84 Impact Factor
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Tanmay Chattopadhyay,
Madhuparna Mukherjee, Arindam Mondal,
Pali Maiti,
Arpita Banerjee,
Kazi Sabnam Banu,
Santanu Bhattacharya,
Bappaditya Roy,
D J Chattopadhyay,
Tapan Kumar Mondal,
Munirathinam Nethaji,
Ennio Zangrando,
Debasis Das
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ABSTRACT: A new dinuclear nickel(II) complex, [Ni(2)(LH(2))(H(2)O)(2)(OH)(NO(3))](NO(3))(3) (1), of an "end-off" compartmental ligand 2,6-bis(N-ethylpiperazine-iminomethyl)-4-methyl-phenolato, has been synthesized and structurally characterized. The X-ray single crystal structure analysis shows that the piperazine moieties assume the expected chair conformation and are protonated. The complex 1 exhibits versatile catalytic activities of biological significance, viz. catecholase, phosphatase, and DNA cleavage activities, etc. The catecholase activity of the complex observed is very dependent on the nature of the solvent. In acetonitrile medium, the complex is inactive to exhibit catecholase activity. On the other hand, in methanol, it catalyzes not only the oxidation of 3,5-di-tert-butylcatechol (3,5-DTBC) but also tetrachlorocatechol (TCC), a catechol which is very difficult to oxidize, under aerobic conditions. UV-vis spectroscopic investigation shows that TCC oxidation proceeds through the formation of an intermediate. The intermediate has been characterized by an electron spray ionizaton-mass spectrometry study, which suggests a bidentate rather than a monodentate mode of TCC coordination in that intermediate, and this proposition have been verified by density functional theory calculation. The complex also exhibits phosphatase (with substrate p-nitrophenylphosphate) and DNA cleavage activities. The DNA cleavage activity exhibited by complex 1 most probably proceeds through a hydroxyl radical pathway. The bioactivity study suggests the possible applications of complex 1 as a site specific recognition of DNA and/or as an anticancer agent.
Inorganic Chemistry 03/2010; 49(7):3121-9. · 4.60 Impact Factor
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ABSTRACT: Chandipura virus, a member of the rhabdoviridae family and vesiculovirus genera, has recently emerged as human pathogen that is associated with a number of outbreaks in different parts of India. Although, the virus closely resembles with the prototype vesiculovirus, Vesicular Stomatitis Virus, it could be readily distinguished by its ability to infect humans. Studies on Chandipura virus while shed light into distinct stages of viral infection; it may also allow us to identify potential drug targets for antiviral therapy. In this review, we have summarized our current understanding of Chandipura virus life cycle at the molecular detail with particular interest in viral RNA metabolisms, namely transcription, replication and packaging of viral RNA into nucleocapsid structure. Contemporary research on otherwise extensively studied family member Vesicular Stomatitis Virus has also been addressed to present a more comprehensive picture of vesiculovirus life cycle. Finally, we reveal examples of protein economy in Chandipura virus life-cycle whereby each viral protein has evolved complexity to perform multiple tasks.
Bioscience Reports 11/2007; 27(4-5):275-98. · 2.38 Impact Factor
