The study of degradation mechanisms of glyco-engineered plant produced anti-rabies monoclonal antibodies E559 and 62-71-3

Rabies is an ancient and neglected zoonotic disease caused by the rabies virus, a neurotropic RNA virus that belongs to the Rhabdoviridae family, genus Lyssavirus. It remains an important public health problem as there are cost and health concerns imposed by the current human post exposure prophylaxis therapy. The use of monoclonal antibodies (mAbs) is therefore an attractive alternative. Rabies mostly affects people that reside in resource-limited areas where there are occasional failures in the cold-chain. These environmental changes may upset the stability of the mAbs. This study focused on mAbs 62-71-3 and E559; their structures, responses to freeze/thaw (F/T) and exposure to reactive oxygen species were therefore studied with the aid of a wide range of biophysical and in silico techniques in order to elucidate their stability and identify aggregation prone regions. E559 was found to be less stable than 62-71-3. The complementarity determining regions (CDR) contributed the most to its instability, more specifically: peptides (EIWD102)-E-99 and (92)ATSPYT(97) found in CDR3, Trp33 found in CDR1 and the oxidised Met34. The constant region "(158)SWNSGALTGHTFPAVL(175)" was also flagged by the special aggregation propensity (SAP) tool and F/T experiments to be highly prone to aggregation. The E559 peptides "(4)LQESGSVL(11) from the heavy chain and (4)LTQSPSSL(11) from the light chain, were also highly affected by F/T. These residues may serve as good candidates for mutation, in the aim to bring forward more stable therapeutic antibodies, thus paving a way to a more safe and efficacious antibody-based cocktail treatment against rabies

Active-Site Mutations in the South African Human Immunodeficiency Virus Type 1 Subtype C Protease Have a Significant Impact on Clinical Inhibitor Binding: Kinetic and Thermodynamic Study▿

Human immunodeficiency virus (HIV) infections in sub-Saharan Africa represent about 56% of global infections. Study of active-site mutations (the V82A single mutation and the V82F I84V double mutation) in the less-studied South African HIV type 1 subtype C (C-SA) protease indicated that neither mutation had a significant impact on the proteolytic functioning of the protease. However, the binding affinities of, and inhibition by, saquinavir, ritonavir, indinavir, and nelfinavir were weaker for each variant than for the wild-type protease, with the double mutant exhibiting the most dramatic change. Therefore, our results show that the C-SA V82F I84V double mutation decreased the binding affinities of protease inhibitors to levels significantly lower than that required for effective inhibition