Biophysical characterisation of the interaction between the S.mansoni G4LZI3 protein and the NAD binding domain of glyceraldehyde-3 phosphate dehydrogenase

A thesis submitted to the Faculty of Science, Agriculture and Engineering in fulfilment of the requirements for the Master of Sciences in the Department of Biochemistry and Microbiology at the University of Zululand, South Africa, 2021.

Schistosomiasis remains a continuous social and economic development burden especially in sub-Saharan Africa where more than 80% of people infected reside. It has been labeled the hotspot of global burden where the most neglected tropical diseases are concentrated. Several countries have, however, made considerable progress in control of the schistosomiasis morbidity through mass drug administration of Praziquantel (PZQ) to school-aged children and other high risk individuals. Although, the extensive use of PZQ as the main drug readily available for large scale treatment of schistosomiasis in most of African regions has raised concerns regarding the emergence of PZQ-resistant schistosome species. The design and development of an alternative drug and/or vaccine against the disease has become an urgent requirement. Hence this study aimed to explore an interaction between two Schistosoma mansoni proteins, G4LZI3 and the NAD binding domain of G3PDH, that are crucial for protection and energy generation of the worm post intra-mammalian invasion. Prior to the in vitro studies, numerous bioinformatics studies were done in order to obtain and examine the conformation of the structures of the two proteins for a preferred alignment prior to interaction. Additionally, the employment of these studies facilitated a better understanding of the physico-chemical properties of the proteins as well as their behavior upon binding and also ‘fishing’ of a potential inhibitor. Molecular docking was employed, which yielded a suitable conformation with minimum binding scores and high binding affinity of the two proteins with the inhibitor (pentalenolactane). The yielded conformation was subjected to molecular dynamics simulation which included RMSD, RMSF and RoG in order to investigate the conformational transformation on the complex Apo-G4LZI3 and smNAD binding domain of G3PDH upon binding of the inhibitor. With the success of the in silico studies, the G4LZI3 protein was recombinantly expressed and purified to homogeneity to iii yield an amount of 12,63mg/ml, which is sufficient to carry out further studies. However, even though the smNAD binding domain protein was successfully over-expressed in bacteria, it was very difficult to achieve complete homogenous purification of this protein thus, downstream in vitro interaction experiments could not be conducted. However, the results obtained from the in silico study provides a comprehensive evidence for future studies based on the design and development of a drug/vaccine for schistosomiasis. Key words: G4LZI3, glyceraldehyde 3-phosphate dehydrogenase, molecular dynamic simulations, Praziquantel, Schistosoma mansoni, .