A novel biosensor for inorganic phosphate (Pi) has been developed based on the phosphate binding protein of Escherichia coli. Two cysteine mutations were introduced and labeled with 6-iodoacetamidotetramethylrhodamine. When physically close to each other and correctly oriented, two rhodamine dyes interact to form a noncovalent dimer. In this state, they have little or no fluorescence, unlike the high fluorescence intensity of monomeric rhodamine. The labeling sites were so placed that the distance and orientation between the rhodamines change as a consequence of the conformational change associated with Pi binding. This movement alters the extent of interaction between the dyes. The best mutant of those tested (A17C, A197C) gives rise on average to approximately 18-fold increase in fluorescence intensity as Pi binds. The kinetics of interaction with Pi were measured at 10 degrees C. Under these conditions, the fluorescence increase associated with Pi binding has a maximum rate of 267 s-1. The Pi dissociation rate is 6.6 s-1, and the dissociation constant is 70 nM. An application of the sensor is demonstrated for measuring ATP hydrolysis in real time as a helicase moves along DNA. Advantages of the new sensor are discussed, both in terms of the use of a rhodamine fluorophore and the potential of this double labeling strategy.

The SH3 domain of Bruton's tyrosine kinase (Btk) is preceded by the Tec homology (TH) region containing prolinerich sequences. We have studied a protein fragment containing both the Btk SH3 domain and the proline-rich sequences of the TH region (PRR-SH3). Intermolecular NMR cross-relaxation measurements, gel permeation chromatography profiles, titrations with proline-rich peptides, and 15 N NMR relaxation measurements are all consistent with a monomer^dimer equilibrium with a dissociation constant on the order of 60 W WM. The intermolecular interactions do, at least in part, involve prolinerich sequences in the TH region. This behavior of Btk PRR-SH3 may have implications for the functional action of Btk. ß

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Several mechanisms are involved in the regulation of cellular signaling. Bruton tyrosine kinase (Btk) of the Tec family contains in the Tec homology (TH) domain a proline-rich region (PRR) capable of interacting with several SH3 domains. The Btk has the SH3 domain adjacent to the TH domain. CD and fluorescence spectroscopy were used to study the binding of two peptides corresponding to segments in the PRR to the Btk SH3 domain. The peptide for the N-terminal half of the PRR binds specifically, whereas the other peptide had hardly any affinity. The TH domain has about four times lower affinity to the SH3 domain than the peptide, 17.0 vs 3.9 microM. The interaction was further tested with an SH3 domain construct that contained the PRR. The two peptides cannot compete for the binding to the extended protein and the TH domain has two times lower affinity to the extended SH3 domain. The intra- or intermolecular interaction between the TH and SH3 domain might have regulatory function also in the other Tec family members.

Introduction: Nigeria has the largest burden of Sickle Cell Disease (SCD) with estimated 100,000 new born affected annually. SCD is a Hemoglobin (Hb) disorder with the major form resulting from the substitution of a polar glutamate (Glu) by non-polar Valine (Val) in an invariant region of Hbβ chain-subunit. Species of Hb found in the sickle cell trait are HbA and HbS in a 60:40 proportion, in SCD only HbS, in the HbC disease only HbC, and in the SC disease it's HbS and HbC in a 50:50 equal proportion. Objective: This paper reviews herbal medicines usage in sub-Saharan Africa (sSA) to ameliorate the crisis associated with SCD. The model Hb tetramer suggests a higher membrane affinity of HbS and HbC, promoting dehydration of RBCs, with concomitant in vivo crystallization. Some drawbacks using these herbal drugs include; poor bioavailability and the lack of proper pharmacovigilance monitoring procedures arising from weak governance structure combined with under reporting of herbal usage to physicians were discussed. Probable epigenetic loci that could be targeted using phytomedicines for effective SCD management were also discussed. Methods: Using search engines, several databases including Google scholar, PubMed, Academic Resource Index were utilized as a source for relevant publications/ literature. The protein coordinates for the Hb tetramer were obtained from the Protein Data Bank (PDB). Conclusion: Manipulation of epigenetics to achieve better SCD management involves careful thinking. Herein, we discuss some epigenetic interactions that could be putatively tweaked with a view of enhancing soluble bioactive small molecular components with the potential to reactivate γ -globin genes, thereby boosting immune response in patient with SCD.

In the rural communities of sub-Saharan African (sSA) countries, malaria is being managed using phytocompounds. Artesunate is reported to inhibit Gephyrin E, a central, multi-domain scaffolding protein of inhibitory post-synapses. Neem plant and its metabolites like azadirachtin are being indicated for management of malaria by traditional healers. The present study was aimed to cheminformatically analyse the binding potential of artesunate and azadirachtin with various reactive moieties of Gephyrin E, to reduce malaria scourge. With molecular dynamics (MD), binding free energy estimation and binding affinity of artesunate and azadirachtin to Gephyrin E was done. GRIP docking was done to study the interactions of these test ligands with Gephyrin E (6FGC). MD simulation gave insights to structural changes upon binding of artesunate and azadirachtin in the ligand-binding pocket of Gephyrin E. Root mean square deviation (RMSD) and root mean square fluctuation (RMSF) were calculated. From the estimation, azadirachtin had a total binding energy of −36.97 kcal/mol; artesunate had a binding energy of −35.73 kcal/mol. The GRIP docking results provided a clearer evidence that artesunate has comparatively better binding affinity to Gephyrin E than azadirachtin, and the critical binding sites (in activity order) were cavity 3, 2, 8, and 6 for artesunate while for azadirachtin, it was cavity 6, 3, 8, and 2. The GRIP docking provided detailed interactions at the atomic levels, providing evidence; both compounds have chances to overcome the drug resistance problem, albeit higher for artesunate. Our findings added another piece of evidence that azadirachtin may be effective as an anti-malarial agent. The results herein may provide impetus for more studies into bioactive components of plant origin towards the effective management of malaria disease phenotype.

The effects of ethylenediaminetetra-acetic acid (EDTA) on germination, length of stem, area of leaf, fresh weight, level of lipid per oxidation, alkaline phosphatase, acid phosphatase, super oxide dismutase (SOD) and catalase in the roots of cadmium (Cd) treated maize (Zea mays L) and cowpea (Vigna unguiculata L) seedlings after 7 and 21 days of germination were determined in this study. The results obtained, indicate that at the end of 7 and 21 days of exposure to Cd, percentage germination of the seeds were not significantly different in both control and test soil (p>0.05). Morphological parameters (area of leaf, length of stem and fresh weight) were significantly reduced by Cd after 7 and 21 days. The supplementation of the soil sample with EDTA (0.5 mM or 1.0 mM) reversed the effect of Cd on these parameters as it significantly increased length of stems, area of leaf and plant fresh weight. There was a significant decrease in root acid phosphatase, root alkaline phosphatase, super-oxide dismutase (SOD) and catalase activity in both plant species. Both used doses of EDTA to ameliorate the above biochemical parameters. Increased level of root lipid peroxidation in Cd treated maize and cowpea seedlings was observed after 7 and 21 days of germination. Albeit, the level of lipid peroxidation in the root of Cd treated maize was significantly higher than that of cowpea, an indication that cowpea may be more tolerant than maize to Cd toxicity. The treatment of plant with, concentrations of EDTA (0.5 and 1.0 mM) failed to decrease the Cd induced, but increased the level of root lipid peroxidation. These results indicate that EDTA (0.5 mM and 1.0 mM) could be used for the treatment of Cd toxicity in plants; although, EDTA did not totally protect cowpea seedling from oxidative stress.

Malaria is a major global health problem with the greatest burden in sub-Saharan Africa (sSA). Unfortunately, Nigeria accounts for 25 percent of the world’s malaria burden and it accounts for more deaths than HIV/AIDS. The causative agent of malaria is plasmodium species. This paper reviews the current approaches to inhibiting plasmodium transmission, and the phyto active compound currently in use in the sSA (particularly in Nigeria) with the goal to ameliorate the high incidence of malaria and to correlating it with recent progress and scientific understanding. Using search engines, several databases including Google scholar, Pub Med, Academic Resource Index, Scopus, etcetera, were utilized to source for relevant publications and literatures. The complex life cycle of the Plasmodium species (causative agent of malaria) gives room for measures that can disrupt its completion. Several methods are currently being tested and experimented on to disrupt the parasite transmission. The disruption of a cell surface transport protein, Feline Leukemia Virus subgroup C Receptor (FLVCR) that pumps heme out of the cell; Gene silencing-techniques used to reduce the levels of FLVCR in the mosquito gut; Prevention of the interaction between the plasmodium TRAP and the Anopheles Saglin protein, which aid the malaria parasite invasion of the mosquito salivary gland; Prevention of the Interaction of Surface Enolase and Plasminogen of Mammalian Blood, disrupting an important role in ookinete invasion of the mosquito midgut; the use of Plants with antimicrobial peptides(cyclotide), that possess structural similarities to SM1 peptide, an inhibitor of plasmodium TRAP-saglin binding;and Use of Phyto-Active Compounds to Block Plasmodium Transmission. These approaches are novel methods in the control and transmission of plasmodium species/malaria. Chemically, phytochemicals with structural similarities to artemisinin, (asesquiteterpene lactone containing an unusual peroxide bridge) is thought of to be present in certain plants with antimalarial and other medicinal value.