Systems biology aims to develop mathematical models of biological systems by integrating experimental and theoretical techniques by leveraging on the genome wide data to unravel the complexity of gene regulation. Despite the availability of effective chemotherapy, leishmaniasis continues to be one of the major parasitic infections that affect the human population worldwide. Currently, little is known of the structural biology of the parasites that are responsible for the disease and few attempts have been made to develop second generation drugs, which may become essential if multi-drug resistance arises. These facts make the discovery of novel drug targets a priority. Multiscale modeling and simulation techniques permit us to study the spatial and temporal properties of large systems to be simulated using atomic-detail structures. The estimation of kinetic parameters for the mathematical modeling provides a basis for iterative manipulation of biochemical pathways. In this paper, emphasis is laid on the discrete regulation of gene or protein expression as modeling can be done based on pure qualitative knowledge about interaction between genes or proteins that is widely available from the existing experimental methodologies. IPC synthase is one of them, believed to play a pivotal role in the pathogenesis of Leishmania sp. and resides in an acidic macrophage phagolysosome, defining a new class of eukaryotic sphingolipid synthases. This work will facilitate the rational development of inhibitors against a protozoan enzyme with no mammalian equivalent, leading to the prospect of anti-protozoal compounds with minimal toxic side effects. Henceforth, it can be said that exploiting the interactome for novel human drug targets could provide new therapeutic avenues towards the treatment of infectious diseases, which could ameliorate the growing clinical challenge of drug-resistant infections.

Cutaneous leishmaniasis affects nearly 0.7 to 1.3 million people annually. Treatment of this disease is difficult due to lack of appropriate medication and the growing problem of drug resistance. Natural compounds such as coumarins serve as complementary therapeutic agents in addition to the current treatment modalities. In this study, we have performed an in-silico screening of the coumarin derivatives and their anti-leishmanial properties has been explored both in-vitro and in-vivo. One of the compounds (compound 2) exhibited leishmanicidal activity and to further study its properties, nanoliposomal formulation of the compound was developed. Treatment of cutaneous lesions in BALB/c mice with compound 2 showed significantly reduced lesion size as compared to the untreated mice (p<0.05) suggesting that compound 2 may possess anti-leishmanial properties.

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BackgroundSelection pressure governs the relative mutability and the conservedness of a protein across the protein family. Biomolecules (DNA, RNA and proteins) continuously evolve under the effect of evolutionary pressure that arises as a consequence of the host parasite interaction. IPCS (Inositol phosphorylceramide synthase), SPL (Sphingosine-1-P lyase) and SPT (Serine palmitoyl transferase) represent three important enzymes involved in the sphingolipid metabolism of Leishmania. These enzymes are responsible for maintaining the viability and infectivity of the parasite and have been classified as druggable targets in the parasite metabolome.ResultsThe present work relates to the role of selection pressure deciding functional conservedness and divergence of the drug targets. IPCS and SPL protein families appear to diverge from the SPT family. The three protein families were largely under the influence of purifying selection and were moderately conserved baring two residues in the IPCS protein which were under the influence of positive selection. To further explore the selection pressure at the codon level, codon usage bias indices were calculated to analyze genes for their synonymous codon usage pattern. IPCS gene exhibited slightly lower codon bias as compared to SPL and SPT protein families.ConclusionEvolutionary tracing of the proposed drug targets has been done with a viewpoint that the amino-acids lining the drug binding pocket should have a lower evolvability. Sites under positive selection (HIS20 and CYS30 of IPCS) should be avoided during devising strategies for inhibitor design.

Leishmania major is the causative agent of cutaneous leishmaniasis which affects over 1 million people in 88 different countries. The incidence of this disease is on the rise due to the current problems associated with the present chemotherapeutics. In addition, Leishmania confronts resistance to the traditional drugs like sodium stibogluconate and newer repurposed drugs like miltefosine. ABC transporters are involved in the development of drug resistance. Miltefosine, the drug used for the treatment of leishmaniasis, is effluxed by P4 ATPase and ABC transporter, which is the prime focus of our study in this paper. P4 ATPase (MDR1) along with an unnamed protein (cdc50) translocates miltefosine from the outer to the inner leaflet by the process of flipping which is ATP driven. In contrast, miltefosine also escapes from the cells by an energy dependent mechanism that involves the ABC transporter protein (ABC). It is known that certain genes in the parasite amplify the portions of a gene which encodes ABC transporter and P4 ATPase involved in translocating phospholipids and hence resistance to miltefosine. We observed the ABC and P4 ATPase genes, 39 T-box elements were observed in the ABC transporter protein and three elements were observed in the P4 ATPase gene suggesting its role in transcription regulation. To the best of our knowledge, there are no structural and regulatory reports on these two proteins in L. major. Computational structural biology tools may aid in understanding the interaction of miltefosine with the P4-ATPase-cdc50 complex and the ABC transporter. This can be achieved by modeling the target protein structures, studying the dynamics associated with the different domains of the protein and later using activators and inhibitors to alter the functioning of the protein. Molecular dynamics simulation with a lipid bilayer is performed to investigate the conformational changes and structure-activity relationship. As transporters are difficult to model, the relevant structural motifs and domains may help to understand the allosteric relation with the substrate and the cofactors. The dynamics of a protein molecule ultimately defines the functional mechanism involving excursions of multiple conformational states. To understand these functional mechanisms of transporter proteins, computational modeling and simulations will be carried out with the goal of elucidating the atomistic details of allosteric conformational transitions and propagations during the transport processes. In particular these studies are designed to investigate the critical structural and dynamic elements that determine individual and combined ligand-binding specificities, the interactions among transporters, their coupled-proteins and the associations of transporters within the lipid bilayer. The nature of results from such studies also makes it possible to rationally optimize existing ligands for these proteins and develop some new compounds that can shift the conformational equilibrium of transporters which may aid in functional stu...

A database that integrates all the information required for biological processing is essential to be stored in one platform. We have attempted to create one such integrated database that can be a one stop shop for the essential features required to fetch valuable result. LmSmdB (L. major and S. mansoni database) is an integrated database that accounts for the biological networks and regulatory pathways computationally determined by integrating the knowledge of the genome sequences of the mentioned organisms. It is the first database of its kind that has together with the network designing showed the simulation pattern of the product. This database intends to create a comprehensive canopy for the regulation of lipid metabolism reaction in the parasite by integrating the transcription factors, regulatory genes and the protein products controlled by the transcription factors and hence operating the metabolism at genetic level.

Inositol phosphorylceramide synthase (IPCS) has emerged as an important, interesting and attractive target in the sphingolipid metabolism of Leishmania. IPCS catalyzes the conversion of ceramide to IPC which forms the most predominant sphingolipid in Leishmania. IPCS has no mammalian equivalent and also plays an important role in maintaining the infectivity and viability of the parasite. The present study explores the possibility of targeting IPCS; development of suitable inhibitors for the same would serve as a treatment strategy for the infectious disease leishmaniasis. Five coumarin derivatives were developed as inhibitors of IPCS protein. Molecular dynamics simulations of the complexes of IPCS with these inhibitors were performed which provided insights into the binding modes of the inhibitors. In vitro screening of the top three compounds has resulted in the identification of one of the compounds (compound 3) which shows little cytotoxic effects. This compound therefore represents a good starting point for further in vivo experimentation and could possibly serve as an important drug candidate for the treatment of leishmaniasis.

Building circuits and studying their behavior in cells is a major goal of systems and synthetic biology. Synthetic biology enables the precise control of cellular states for systems studies, the discovery of novel parts, control strategies, and interactions for the design of robust synthetic systems. To the best of our knowledge, there are no literature reports for the synthetic circuit construction for protozoan parasites. This paper describes the construction of genetic circuit for the targeted enzyme inositol phosphorylceramide synthase belonging to the protozoan parasite Leishmania. To explore the dynamic nature of the circuit designed, simulation was done followed by circuit validation by qualitative and quantitative approaches. The genetic circuit designed for inositol phosphorylceramide synthase (Biomodels Database-MODEL1208030000) shows responsiveness, oscillatory and bistable behavior, together with intrinsic robustness.