Drug targets

Target discovery using the molecular approach, as opposed to the more traditional systems approach
requires the study of the cellular or biological process underlying a condition or disease. The approaches
that are employed by the “bench” scientist may be genetic, genomic or proteomic and each has its rightful place
in the drug-target discovery process. Affinity-based proteomic techniques currently used in drug-discovery draw upon
several disciplines, synthetic chemistry, cell-biology, biochemistry and mass spectrometry. An important component of
such techniques is the probe that is specifically designed to pick out a protein or set of proteins from amongst the varied
thousands in a cell lysate. A second component, that is just as important, is liquid-chromatography tandem massspectrometry
(LC-MS/MS). LC-MS/MS and the supporting theoretical framework has come of age and is the tool of
choice for protein identification and quantification. These proteomic tools are critical to maintaining the drug-candidate
supply, in the larger context of drug discovery.

Heat shock proteins (Hsps) are ubiquitous and
conserved molecules whose main function is to facilitate
protein folding (molecular chaperone function). Some
Hsps such as Hsp70 are also involved in protein folding,
protein trafficking and ubiquitination of misfolded
protein to facilitate their degradation. Hsps occur in functional networks and
hence their coordinated expression is important in
proteostasis. Some Hsps are stress-induced while others
are expressed constitutively. Because of their role as
sentinels of protein folding, it is not surprising that some
Hsps are essential and for this reason, they constitute
potential drug target. However, their high conservation is a snag to their
prospects as drug targets. In spite of this, drugs
targeting some Hsps especially Hsp90 show great
potential and some of them have entered clinical trials
for cancer treatment. In addition,
there are prospects to target Hsps in treatment of
infectious diseases such as malaria

The immune system protects a host from foreign pathogens. In rare cases, the immune system can attack the cells of the host organism causing autoimmune diseases. We outline a computational framework that combines bioinformatics and network analysis with an emerging targets platform. The computational framework presented here can be used to find drug targets for autoimmune diseases. It can also be used to find existing drugs that can be repurposed to treat autoimmune diseases based on networks of interactions or similarities between different diseases. Information on which gene regions are associated with the disease (single nucleotide polymorphisms) can be used in gene therapy when that technique becomes viable. Our analysis also revealed immune cell subtypes that are implicated in these diseases. These immune cell subtypes can be selected for immunotherapy experiments. Finally, our analysis also reveals intra-cellular and protein-protein interaction networks and pathways that can be targeted with small molecule inhibitors. The downstream off-target effects of these inhibitors can also be determined from such a network analysis. In summary, our computational framework can be used to find novel therapeutics for autoimmune diseases and potentially even other dysfunctions.

Background
Infections caused by Salmonella enterica, a Gram-negative facultative anaerobic bacteria belonging to the family of Enterobacteriaceae, are major threats to the health of humans and animals. The recent availability of complete genome data of pathogenic strains of the S. enterica gives new avenues for the identification of drug targets and drug candidates. We have used the genomic and metabolic pathway data to identify pathways and proteins essential to the pathogen and absent from the host.
Methods
We took the whole proteome sequence data of 42 strains of S. enterica and Homo sapiens along with KEGG-annotated metabolic pathway data, clustered proteins sequences using CD-HIT, identified essential genes using DEG database and discarded S. enterica homologs of human proteins in unique metabolic pathways (UMPs) and characterized hypothetical proteins with SVM-prot and InterProScan. Through this core proteomic analysis we have identified enzymes essential to the pathogen.
Results
The identification of 73 enzymes common in 42 strains of S. enterica is the real strength of the current study.We proposed all 73 unexplored enzymes as potential drug targets against the infections caused by the S. enterica. The study is comprehensive around S. enterica and simultaneously considered every possible pathogenic strain of S. enterica. This comprehensiveness turned the current study significant since, to the best of our knowledge it is the first subtractive core proteomic analysis of the unique metabolic pathways applied to any pathogen for the identification of drug targets. We applied extensive computational methods to shortlist few potential drug targets considering the druggability criteria e.g. Non-homologous to the human host, essential to the pathogen and playing significant role in essential metabolic pathways of the pathogen (i.e. S. enterica). In the current study, the subtractive proteomics through a novel approach was applied i.e. by considering only proteins of the unique metabolic pathways of the pathogens and mining the proteomic data of all completely sequenced strains of the pathogen, thus improving the quality and application of the results. We believe that the sharing of the knowledge from this study would eventually lead to bring about novel and unique therapeutic regimens against the infections caused by the S. enterica.

The function of a protein is dependent on its three-dimensional
structure. However, numerous proteins lacking intrinsic globular 3D structure under physiological
conditions had been recognized. These proteins are frequently involved in some of the most critical
cellular control mechanisms and it appears that their rapid turnover, aided by their unstructured nature
in the unbound state, provides a level of control that allows rapid and accurate responses of the cell to
changing environmental conditions. Approach: A significant number of proteins known to be
involved in protein deposition disorders were now considered to Be Intrinsically Unstructured Proteins
(IUPs). For example, Aβ peptide and tau protein in Alzheimer’s disease, PrP in Prion’s disease and α-
Synuclein in Parkinson’s disease. The disorder of intrinsically unstructured proteins (IUP's) was
crucial to their functions. They may adopt defined but extended structures when bound to cognate
ligands. Their amino acid compositions were less hydrophobic than those of soluble proteins. They
lack hydrophobic cores and hence did not become insoluble when heated. About 40% of eukaryotic
proteins had at least one long (>50 residues) disordered region. Roughly 10% of proteins in various
genomes had been predicted to be fully disordered. Presently over 100 IUP's had been identified; none
are enzymes. Obviously, IUP's were greatly underrepresented in the Protein Data Bank, although there
were few cases of an IUP bound to a folded (intrinsically structured) protein. Results: The five
functional categories for intrinsically unstructured proteins and domains were entropic chains (bristles
to ensure spacing, springs, flexible spacers/linkers), effectors (inhibitors and disassemblers),
scavengers, assemblers and display sites. These IUPs could serve as potential targets for Structure
Based Drug Design (SBDD) which stress on the transition from disordered to ordered confirmation
through drug stimulation. Recently an unstructured domain of a regulatory protein had been found to
be involved in inhibiting catalytic activity of insulin receptor and targeting this IUP would provide a
new approach which can be employed in modifying insulin signaling in treatment of diabetes. IUPs
were also involved diseases and disorders such as cardio vascular diseases, cancers and autoimmune
disorders. Unstructured proteins had also been shown to be important components of invasion, survival
and disguising strategies of pathogens such as Plasmodium falciparum. Conclusion: New greater
focuses on proteins that were in some way unstructured normally would promise to provide a greater
understanding of protein function particularly with respect to protein-protein interactions and hence
can give new potential targets for future strategies.

Honey has antioxidant, immunostimulant, antibacterial, and antileishmanial activities. In this study, we evaluated the in vitro babesicidal and theilericidal effects of honey on Babesia bovis, Babesia bigemina, Babesia divergens, Babesia caballi, and Theileria equi. There was noteworthy suppression of growth at a concentration of 0.5% (V/V) for B. bovis, B. bigemina, B. divergens, and T. equi and 1% (V/V) for B. caballi. The IC 50 values of honey were 1.98, 1.82, 0.42, 1.7, and 1.43% (V/V) for B. bovis, B. bigemina, B. divergens, B. caballi, and T. equi, respectively. The growth was entirely repressed at 1% (V/V) for B. divergens, 2.5% (V/V) for T. equi, and 5% (V/V) for B. bovis, B. bigemina, and B. caballi. The regrowth was repressed in the viability test at a concentration of 1% (V/V) for B. divergens, 2.5% (V/V) for T. equi, and 5% (V/V) for B. caballi, B. bovis, and B. bigemina. These results indicate honey as a natural killer of Babesia species and T. equi. Its use in the treatment of clinical cases requires further in vivo evaluation.

Caseinolytic chaperones and proteases (Clp) belong to the AAA1 protein superfamily and
are part of the protein quality control machinery in cells. The eukaryotic parasite Plasmodium
falciparum, the causative agent of malaria, has evolved an elaborate network of Clp proteins
including two distinct ClpB ATPases. ClpB1 and ClpB2 are involved in different aspects of parasitic
proteostasis. ClpB1 is present in the apicoplast, a parasite-specific and plastid-like organelle
hosting various metabolic pathways necessary for parasite growth. ClpB2 localizes to the parasitophorous
vacuole membrane where it drives protein export as core subunit of a parasite-derived
protein secretion complex, the Plasmodium Translocon of Exported proteins (PTEX); this process
is central to parasite virulence and survival in the human host. The functional associations of these
two chaperones with parasite-specific metabolism and protein secretion make them prime drug
targets. ClpB proteins function as unfoldases and disaggregases and share a common architecture
consisting of four domains—a variable N-terminal domain that binds different protein substrates,
followed by two highly conserved catalytic ATPase domains, and a C-terminal domain. Here, we
report and compare the first crystal structures of the N terminal domains of ClpB1 and ClpB2 from
Plasmodium and analyze their molecular surfaces. Solution scattering analysis of the N domain of
ClpB2 shows that the average solution conformation is similar to the crystalline structure. These structures represent the first step towards the characterization of these two malarial chaperones
and the reconstitution of the entire PTEX to aid structure-based design of novel anti-malarial
drugs.