Human serum albumin as a potential drug delivery system for N-methylated hot spot insulin analogs inhibiting hormone aggregation

Abstract

The purpose of this study was to investigate whether Human Serum Albumin (HSA) can bind N-methylated analogs of hot spots of native insulin. Three N-methylated derivatives of the A13–A19 fragment of native insulin were used: L(N-Me)YQLENY (1), LYQ(N-Me)LENY (2), and L(N-Me)YQ(N-Me)LENY (3). The studied N-methylated insulin fragments possess inhibiting potential against hormone aggregation. A variety of research techniques, including spectroscopic methods and microscopy assays, were used to study the interaction of HSA with the N-methylated insulin fragments. Based on spectroscopic measurements with Congo Red and Thioflavin T, all the analyzed N-methylated peptides were able to interact with the HSA surface. The CD spectrum registered for HSA in the presence of L(N-Me)YQLENY showed the smallest content of α-helix conformation, indicating the most compact HSA structure. Based on the results of MST, the dissociation constants (Kd) for complexes of HSA and peptides 1–3 were 19.2 nM (complex 1), 15.6 nM (complex 2), and 8.07 nM (complex 3). Microscopy assays, dynamic light scattering measurements as well as computer simulation of protein–ligand interaction also confirmed the possibility of docking the N-methylated inhibitors within HSA.