Socrates is Alive and Well! The Case for Dialogue and Critical Thinking In Values and Ethics Education

This paper underlines the need for teaching morals and values through critical reflection and active genuine dialogue. It promotes the pedagogy of dialogue within educational institutions, the creation of multi-dimensional learning environments for the cultivation and dissemination of intersubjective understandings of diverse moral world views, the use of critical thinking skills and intellectual traits of mind for ethical decision making, and the communication of values and morals through dialogue. An argument is advanced to show how reflective dialogue lays the groundwork for the creation of initial objective relations in the classroom and forms the basis for the pragmatic implementation of an interpersonal connection characterized by feelings of tolerance, empathy, and respect for the dignity of human beings and their way of life

Biophysical and structural studies of novel F420-dependent oxidoreductases in Mycobacterium tuberculosis

The Mycobacterium tuberculosis (Mtb) genome encodes hundreds of proteins with unknown or putative function based on sequence similarity to characterised proteins. Incomplete knowledge of Mtb functional genomics undermines efforts to understand the complex biological repertoire of this deadly human pathogen. While bioinformatics provides useful clues about gene function, biochemical characterisation of gene products remains essential to accurate annotation, particularly in cases where functional redundancy is suggested based on the presence of multiple homologous proteins. The Mtb genome includes 7 genes which encode conserved hypothetical proteins with unknown function that are annotated as “pyridoxine 5’-phosphate (PNPOx)-like” proteins based on structural similarity: Rv2607, Rv2991, Rv1155, Rv2074, Rv3369c, Rv1875, Rv0121c. PNPOx is a flavin mononucleotide (FMN)-dependent enzyme involved in the biosynthesis of the crucial cofactor, pyridoxal 5’- phosphate (PLP). The work presented in this thesis shows that while Rv2607 exhibits canonical PNPOx activity, Rv1155 and Rv2991 do not have affinity for FMN as do other members of the PNPOx class. They instead bind tightly to the unusual flavin coenzyme F420 by isothermal calorimetry (ITC), saturation transfer difference (STD) NMR, and X-ray crystallography. Although recent studies have drawn attention to F420 due to its role in activating the promising anti-tuberculosis drug PA-824, little is known about the function of this coenzyme in Mtb metabolism. In order to identify the substrate recognition properties of Rv1155 and Rv2991 respectively, a novel application of fragment-based approaches was employed. A library of fragments was designed based on known ligands to flavoenzymes and screened for binding to Rv1155 and Rv2991 in the presence and absence of F420 using STD NMR and differential scanning fluorimetry. Rv1155 and Rv2991 show affinity for distinct classes of fragments. Using a computational method known as “virtual fragment linking,” fragment binding data was used to predict and test potential substrates. This approach to probing structure-function relationships may serve as a generalisable method for exploring functional diversity within a structural class of proteins.This work was supported by the National Institutes of Health-Oxford-Cambridge Scholars Progra

Applications of isothermal titration calorimetry - the research and technical developments from 2011 to 2015

Isothermal titration calorimetry is a widely used biophysical technique for studying the formation or dissociation of molecular complexes. Over the last 5years, much work has been published on the interpretation of isothermal titration calorimetry (ITC) data for single binding and multiple binding sites. As over 80% of ITC papers are on macromolecules of biological origin, this interpretation is challenging. Some researchers have attempted to link the thermodynamics constants to events at the molecular level. This review highlights work carried out using binding sites characterized using x-ray crystallography techniques that allow speculation about individual bond formation and the displacement of individual water molecules during ligand binding and link these events to the thermodynamic constants for binding. The review also considers research conducted with synthetic binding partners where specific binding events like anion-π and π-π interactions were studied. The revival of assays that enable both thermodynamic and kinetic information to be collected from ITC data is highlighted. Lastly, published criticism of ITC research from a physical chemistry perspective is appraised and practical advice provided for researchers unfamiliar with thermodynamics and its interpretation