Doctor of Philosophy, Ludwig Maximilian Universitat Munchen (2010)
Venkata Raveendra Pothineni. "United States Patent US 621079,537 INHIBITORS OF BORRELIA METAL TRANSPORTER FOR TREATMENT OF LYME DISEASE", stanford, Nov 28, 2014
Current Research and Scholarly Interests
high throughput screening new drugs for borrelia burgdorferi
Borreliacidal activity of Borrelia metal transporter A (BmtA) binding small molecules by manganese transport inhibition
DRUG DESIGN DEVELOPMENT AND THERAPY
2015; 9: 805-815
Borrelia burgdorferi, the causative agent of Lyme disease, utilizes manganese (Mn) for its various metabolic needs. We hypothesized that blocking Mn transporter could be a possible approach to inhibit metabolic activity of this pathogen and eliminate the infection. We used a combination of in silico protein structure prediction together with molecular docking to target the Borrelia metal transporter A (BmtA), a single known Mn transporter in Borrelia and screened libraries of FDA approved compounds that could potentially bind to the predicted BmtA structure with high affinity. Tricyclic antihistamines such as loratadine, desloratadine, and 3-hydroxydesloratadine as well as yohimbine and tadalafil demonstrated a tight binding to the in silico folded BmtA transporter. We, then, tested borreliacidal activity and dose response of the shortlisted compounds from this screen using a series of in vitro assays. Amongst the probed compounds, desloratadine exhibited potent borreliacidal activity in vitro at and above 78 μg/mL (250 μM). Borrelia treated with lethal doses of desloratadine exhibited a significant loss of intracellular Mn specifically and a severe structural damage to the bacterial cell wall. Our results support the possibility of developing a novel, targeted therapy to treat Lyme disease by targeting specific metabolic needs of Borrelia.
View details for DOI 10.2147/DDDT.S77063
View details for Web of Science ID 000349239100003
View details for PubMedID 25709405
Comprehensive Analysis of Varicella-Zoster Virus Proteins Using a New Monoclonal Antibody Collection
JOURNAL OF VIROLOGY
2013; 87 (12): 6943-6954
Varicella-zoster virus (VZV) is the etiological agent of chickenpox and shingles. Due to the virus's restricted host and cell type tropism and the lack of tools for VZV proteomics, it is one of the least-characterized human herpesviruses. We generated 251 monoclonal antibodies (MAbs) against 59 of the 71 (83%) currently known unique VZV proteins to characterize VZV protein expression in vitro and in situ. Using this new set of MAbs, 44 viral proteins were detected by Western blotting (WB) and indirect immunofluorescence (IF); 13 were detected by WB only, and 2 were detected by IF only. A large proportion of viral proteins was analyzed for the first time in the context of virus infection. Our study revealed the subcellular localization of 46 proteins, 14 of which were analyzed in detail by confocal microscopy. Seven viral proteins were analyzed in time course experiments and showed a cascade-like temporal gene expression pattern similar to those of other herpesviruses. Furthermore, selected MAbs tested positive on human skin lesions by using immunohistochemistry, demonstrating the wide applicability of the MAb collection. Finally, a significant portion of the VZV-specific antibodies reacted with orthologs of simian varicella virus (SVV), thus enabling the systematic analysis of varicella in a nonhuman primate model system. In summary, this study provides insight into the potential function of numerous VZV proteins and novel tools to systematically study VZV and SVV pathogenesis.
View details for DOI 10.1128/JVI.00407-13
View details for Web of Science ID 000319508600038
View details for PubMedID 23596286
Varicella Zoster Virus ORF25 Gene Product: An Essential Hub Protein Linking Encapsidation Proteins and the Nuclear Egress Complex
JOURNAL OF PROTEOME RESEARCH
2011; 10 (12): 5374-5382
Varicella zoster virus (VZV) ORF25 is a 156 amino acid protein belonging to the approximately 40 core proteins that are conserved throughout the Herpesviridae. By analogy to its functional orthologue UL33 in Herpes simplex virus 1 (HSV-1), ORF25 is thought to be a component of the terminase complex. To investigate how cleavage and encapsidation of viral DNA links to the nuclear egress of mature capsids in VZV, we tested 10 VZV proteins that are predicted to be involved in either of the two processes for protein interactions against each other using three independent protein-protein interaction (PPI) detection systems: the yeast-two-hybrid (Y2H) system, a luminescence based MBP pull-down interaction screening assay (LuMPIS), and a bioluminescence resonance energy transfer (BRET) assay. A set of 20 interactions was consistently detected by at least 2 methods and resulted in a dense interaction network between proteins associated in encapsidation and nuclear egress. The results indicate that the terminase complex in VZV consists of ORF25, ORF30, and ORF45/42 and support a model in which both processes are closely linked to each other. Consistent with its role as a central hub for protein interactions, ORF25 is shown to be essential for VZV replication.
View details for DOI 10.1021/pr200628s
View details for Web of Science ID 000297537200010
View details for PubMedID 21988664
Systematic analysis of the IgG antibody immune response against varicella zoster virus (VZV) using a self-assembled protein microarray
2010; 6 (9): 1604-1610
Varicella zoster virus (VZV) is a human herpesvirus encoding at least 69 distinct viral proteins which causes chickenpox after primary infection and shingles during reactivation and which is particularly important in pregnancy and immunocompromised patients. Current serodiagnostic tests are either based on whole cell lysates or glycoprotein preparations. In order to investigate the humoral immune response to VZV infection or vaccination in more detail, and to improve the currently available diagnostic assays, we developed a nucleic acid programmable protein array (NAPPA) containing all 69 VZV proteins and performed a detailed analysis of 68 sera from individuals with either no, a previous or an acute VZV infection. In addition to the known reactive glycoprotein antigens (ORF 5, ORF 14, ORF 31, ORF 37, ORF 68), we discovered IgG antibodies against a variety of other membrane (ORF 2, ORF 24), capsid (ORF 20, ORF 23, ORF 43) and tegument (ORF 53, ORF 9, ORF 11) proteins, as well as other proteins involved in virus replication and assembly (ORF 25, ORF 26, ORF 28) and the transactivator proteins ORF 12, ORF 62 and ORF 63. All of these antigens were only reactive in a subset of VZV-positive individuals. A subset of the newly identified VZV antigens was validated by western blot analysis. Using these seroreactive new VZV antigens, more sensitive assays and tests distinguishing between different clinical entities may be developed.
View details for DOI 10.1039/c003798b
View details for Web of Science ID 000280868000014
View details for PubMedID 20514382
Improving the yeast two-hybrid system with permutated fusions proteins: the Varicella Zoster Virus interactome.
2010; 8: 8-?
Yeast two-hybrid (Y2H) screens have been among the most powerful methods to detect and analyze protein-protein interactions. However, they suffer from a significant degree of false negatives, i.e. true interactions that are not detected, and to a certain degree from false positives, i.e. interactions that appear to take place only in the context of the Y2H assay. While the fraction of false positives remains difficult to estimate, the fraction of false negatives in typical Y2H screens is on the order of 70-90%. Here we present novel Y2H vectors that significantly decrease the number of false negatives and help to mitigate the false positive problem.We have constructed two new vectors (pGBKCg and pGADCg) that allow us to make both C-terminal fusion proteins of DNA-binding and activation domains. Both vectors can be combined with existing vectors for N-terminal fusions and thus allow four different bait-prey combinations: NN, CC, NC, and CN. We have tested all approximately 4,900 pairwise combinations of the 70 Varicella-Zoster-Virus (VZV) proteins for interactions, using all possible combinations. About approximately 20,000 individual Y2H tests resulted in 182 NN, 89 NC, 149 CN, and 144 CC interactions. Overlap between screens ranged from 17% (NC-CN) to 43% (CN-CC). Performing four screens (i.e. permutations) instead of one resulted in about twice as many interactions and thus much fewer false negatives. In addition, interactions that are found in multiple combinations confirm each other and thus provide a quality score. This study is the first systematic analysis of such N- and C-terminal Y2H vectors.Permutations of C- and N-terminal Y2H vectors dramatically increase the coverage of interactome studies and thus significantly reduce the number of false negatives. We suggest that future interaction screens should use such vector combinations on a routine basis, not the least because they provide a built-in quality score for Y2H interactions that can provide a measure of reproducibility without additional assays.
View details for DOI 10.1186/1477-5956-8-8
View details for PubMedID 20205919