Insulin aggregation kinetics monitored by luminescent probe cis-[Ru(phen)2(ImAC)]+.

• Grant number: 21/11952-9
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Start date: January 01, 2022
• End date: November 30, 2022
• Field of knowledge: Physical Sciences and Mathematics - Chemistry - Inorganic Chemistry
• Principal Investigator: Rose Maria Carlos
• Grantee: Bárbara Patrícia Neves Silva
• Host Institution: Centro de Ciências Exatas e de Tecnologia (CCET). Universidade Federal de São Carlos (UFSCAR). São Carlos , SP, Brazil

Abstract

Diabetes is a chronic progressive disease characterized by high blood glucose levels and associated with tissue and organ damage, therefore, access to affordable treatment, including insulin, is critical to the survival of people with diabetes. Insulin was the first biopharmaceutical produced on an industrial scale for the treatment of insulin-dependent diabetes. Insulin is an amyloid protein that in monomeric form undergoes a self-aggregation process, generating amyloid fibrils. Amyloid protein aggregation is one of the most common, problematic processes found in all stages of development, storage, transport, and administration of biopharmaceuticals because in the form of aggregates it is not bioavailable and any degree of amyloid formation during its manufacture reduces absorption, altering the effectiveness of the treatment. Therefore, knowledge of insulin fibrillation mechanisms is important both to improve pharmaceutical formulations used in the treatment of diabetic patients, as well as to contribute to the knowledge of the molecular characteristics of the formation of amyloid proteins. Recent studies from our laboratory have shown that the luminescent complex cis-[Ru(phen)2(3,4Apy)]2+, (RuApy) 3,4Apy=3,4-aminopyridine, is a luminescent probe sensitive to fibrillar insulin with a detection limit of 0.85 ¼M and a binding affinity of 12.40 ¼M. The fibrillation process of amyloid proteins and insulin, in particular, is dependent on non-covalent intermolecular interactions such as hydrogen bonds, electrostatic interactions as well as hydrophobic contacts. These characteristics motivated us to investigate the insulin aggregation process in real-time by changes in the luminescent properties of the complex [Ru(phen)2(ImAC)]+, (RuImAC), ImAC= 4-imidazole carboxylic acid. This new luminescent probe despite being similar in molecular radius to RuApy differs in charge, and it does not contribute to hydrogen bonding, thus being able to present significant differences in interactions with insulin and altering the aggregation process. We expect that the differences (or not) in insulin aggregation kinetics by RuApy and RuImAC may contribute to the knowledge of the effect and influence of electrostatic interactions and hydrogen bonding in the insulin aggregation process. Our strategy for this new luminescent probe was to promote a tris-bidentate structure in the Ru(II) complex to ensure intense absorption and emission of MLCT (Ru,dpp*,phen) and which hinders the photodissociation of the imidazole ligand. The studies will be carried out using steady-state absorption and emission spectroscopic techniques (quantum emission intensity and yield) and time-resolved (emission lifetime) and fluorescence microscopy imaging techniques.(AU)