Research Articles (Biochemistry)
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- ItemSupporting data on characterisation of linker switch mutants of Plasmodium falciparum heat shock protein 110 and canonical Hsp70(Elsevier Inc., 2021-05-29) Chakafana, Graham; Mudau, Pertunia T.; Zininga, Tawanda; Shonhai, AddmoreHere, we present data on characterisation of the linker of Plasmodium falciparum Hsp110 (PfHsp70-z) relative to the linker of canonical Hsp70s in support of a co-published article [1]. The linker of PfHsp70-z was switched with that of canonical Hsp70s, represented by PfHsp70–1 (cytosolic counterpart of PfHsp70-z) and E. coli Hsp70/DnaK. The datasets represent comparative analyses of PfHsp70-z, PfHsp70–1, and E. coli DnaK, relative to their linker switch mutants; PfHsp70-zLS, PfHsp70–1LS, DnaKLS, respectively. Intrinsic and extrinsic fluorescence spectroscopic analyses were employed to elucidate effects of the mutations on the structural features of the proteins. The structural conformations of the proteins were analysed in the absence as well as presence of nucleotides. In addition, stability of the proteins to stress (pH changes and urea) was also determined. Surface plasmon resonance (SPR) was employed to determine affinity of the proteins for ATP. The relative affinities of PfHsp70-z and PfHsp70–1 for the parasite cytosol localised, J domain co-chaperone, PfHsp40, was determined by SPR analysis. The effect of the linker of PfHsp70-z on the interaction of DnaKLS with DnaJ (a co-chaperone of DnaK), was similarly determined. These data could be used for future investigations involving protein-protein/ligand interactions as described in [1]. The raw data obtained using the various techniques here described are hosted in the Mendeley Data repository at [2].
- ItemComparative characterization of plasmodium falciparum Hsp70-1 relative to E. coli DnaK reveals the functional specificity of the parasite chaperone(Multidisciplinary Digital Publishing Institute (MDPI), 2020-06-04) Lebepe, Charity Mekgwa; Matambanadzo, Pearl Rutendo; Makhoba, Xolani Henry; Achilonu, Ikechukwu; Zininga, Tawanda; Shonhai, AddmoreHsp70 is a conserved molecular chaperone. How Hsp70 exhibits specialized functions across species remains to be understood. Plasmodium falciparum Hsp70-1 (PfHsp70-1) and Escherichia coli DnaK are cytosol localized molecular chaperones that are important for the survival of these two organisms. In the current study, we investigated comparative structure-function features of PfHsp70-1 relative to DnaK and a chimeric protein, KPf, constituted by the ATPase domain of DnaK and the substrate binding domain (SBD) of PfHsp70-1. Recombinant forms of the three Hsp70s exhibited similar secondary and tertiary structural folds. However, compared to DnaK, both KPf and PfHsp70-1 were more stable to heat stress and exhibited higher basal ATPase activity. In addition, PfHsp70-1 preferentially bound to asparagine rich peptide substrates, as opposed to DnaK. Recombinant P. falciparum adenosylmethionine decarboxylase (PfAdoMetDC) co-expressed in E. coli with either KPf or PfHsp70-1 was produced as a fully folded product. Co-expression of PfAdoMetDC with heterologous DnaK in E. coli did not promote folding of the former. However, a combination of supplementary GroEL plus DnaK improved folding of PfAdoMetDC. These findings demonstrated that the SBD of PfHsp70-1 regulates several functional features of the protein and that this molecular chaperone is tailored to facilitate folding of plasmodial proteins.
- ItemThe carbon switch at the level of pyruvate and phosphoenolpyruvate in sulfolobus solfataricus P2(Frontiers Media, 2019-04-12) Haferkamp, Patrick; Tjaden, Britta; Shen, Lu; Brasen, Christopher; Kouril, Theresa; Siebers, BettinaSulfolobus solfataricus P2 grows on different carbohydrates as well as alcohols, peptides and amino acids. Carbohydrates such as D-glucose or D-galactose are degraded via the modified, branched Entner–Doudoroff (ED) pathway whereas growth on peptides requires the Embden–Meyerhof–Parnas (EMP) pathway for gluconeogenesis. As for most hyperthermophilic Archaea an important control point is established at the level of triosephophate conversion, however, the regulation at the level of pyruvate/phosphoenolpyruvate conversion was not tackled so far. Here we describe the cloning, expression, purification and characterization of the pyruvate kinase (PK, SSO0981) and the phosphoenolpyruvate synthetase (PEPS, SSO0883) of Sul. solfataricus. The PK showed only catabolic activity [catalytic efficiency (PEP): 627.95 mM⁻¹s⁻¹, 70°C] with phosphoenolpyruvate as substrate and ADP as phosphate acceptor and was allosterically inhibited by ATP and isocitrate (Ki 0.8 mM). The PEPS was reversible, however, exhibited preferred activity in the gluconeogenic direction [catalytic efficiency (pyruvate): 1.04 mM⁻¹s⁻¹, 70°C] and showed some inhibition by AMP and α-ketoglutarate. The gene SSO2829 annotated as PEPS/pyruvate:phosphate dikinase (PPDK) revealed neither PEPS nor PPDK activity. Our studies suggest that the energy charge of the cell as well as the availability of building blocks in the citric acid cycle and the carbon/nitrogen balance plays a major role in the Sul. solfataricus carbon switch. The comparison of regulatory features of well-studied hyperthermophilic Archaea reveals a close link and sophisticated coordination between the respective sugar kinases and the kinetic and regulatory properties of the enzymes at the level of PEP-pyruvate conversion.
- ItemUncovering the effects of heterogeneity and parameter sensitivity on within‑host dynamics of disease : malaria as a case study(BMC (part of Springer Nature), 2021-07-24) Horn, Shade; Snoep, Jacky L.; Van Niekerk, David D.Background: The fidelity and reliability of disease model predictions depend on accurate and precise descriptions of processes and determination of parameters. Various models exist to describe within-host dynamics during malaria infection but there is a shortage of clinical data that can be used to quantitatively validate them and establish confidence in their predictions. In addition, model parameters often contain a degree of uncertainty and show variations between individuals, potentially undermining the reliability of model predictions. In this study models were reproduced and analysed by means of robustness, uncertainty, local sensitivity and local sensitivity robustness analysis to establish confidence in their predictions. Results: Components of the immune system are responsible for the most uncertainty in model outputs, while disease associated variables showed the greatest sensitivity for these components. All models showed a comparable degree of robustness but displayed different ranges in their predictions. In these different ranges, sensitivities were well-preserved in three of the four models. Conclusion: Analyses of the effects of parameter variations in models can provide a comparative tool for the evaluation of model predictions. In addition, it can assist in uncovering model weak points and, in the case of disease models, be used to identify possible points for therapeutic intervention.
- ItemHarmonizing semantic annotations for computational models in biology(Oxford University Press, 2019) Neal, Maxwell Lewis; Konig, Matthias; Nickerson, David; Mısırlı, Goksel; Kalbasi, Reza; Drager, Andreas; Atalag, Koray; Chelliah, Vijayalakshmi; Cooling, Michael T.; Cook, Daniel L.; Crook, Sharon; De Alba, Miguel; Friedman, Samuel H.; Garny, Alan; Gennari, John H.; Gleeson, Padraig; Golebiewski, Martin; Hucka, Michael; Juty, Nick; Myers, Chris; Olivier, Brett G.; Sauro, Herbert M.; Scharm, Martin; Snoep, Jacky L.; Toure, Vasundra; Wipat, Anil; Wolkenhauer, Olaf; Waltemath, DagmarLife science researchers use computational models to articulate and test hypotheses about the behavior of biological systems. Semantic annotation is a critical component for enhancing the interoperability and reusability of such models as well as for the integration of the data needed for model parameterization and validation. Encoded as machine-readable links to knowledge resource terms, semantic annotations describe the computational or biological meaning of what models and data represent. These annotations help researchers find and repurpose models, accelerate model composition and enable knowledge integration across model repositories and experimental data stores. However, realizing the potential benefits of semantic annotation requires the development of model annotation standards that adhere to a community-based annotation protocol.Without such standards, tool developers must account for a variety of annotation formats and approaches, a situation that can become prohibitively cumbersome and which can defeat the purpose of linking model elements to controlled knowledge resource terms. Currently, no consensus protocol for semantic annotation exists among the larger biological modeling community. Here, we report on the landscape of current annotation practices among the COmputational Modeling in BIology NEtwork community and provide a set of recommendations for building a consensus approach to semantic annotation.