Abstract
Apolipoprotein A-IV (apoA-IV) is an abundant lipid-binding protein in blood plasma. We previously reported that apoA-IV, as an endogenous inhibitor, competitively binds platelet αIIbβ3 integrin from its N-terminal residues, reducing the potential risk of thrombosis. This study aims to investigate how the apoA-IVQ360H and apoA-IVT347S mutations affect the structure and function of apoA-IV. These mutations are linked to increased risk of cardiovascular diseases because of multiple single-nucleotide polymorphisms in the C-terminal region of apoA-IV. We postulate that the structural hindrance caused by the C-terminal motifs may impede the binding of apoA-IV to platelets at its N-terminal binding site. However, the mechanistic impact of Q360H and T347S polymorphisms on this intermolecular interaction and their potential contribution to the development of cardiovascular disease have not been adequately investigated. To address this, recombinant forms of human apoA-IVWT, apoA-IVQ360H, and apoA-IVT347S variants were produced, and the structural stability, dimerization, and molecular dynamics of the C terminus were examined utilizing biophysical techniques, including fluorescence anisotropy, fluorescence spectrophotometry, circular dichroism, and biolayer interferometry methods. Our results showed a decreased fraction of α-helix structure in apoA-IVQ360H and apoA-IVT347S compared with the WT, and the inhibitory effect of dimerized apoA-IV on platelet aggregation was reduced in apoA-IVQ360H and apoA-IVT347S variants. Binding kinetics of examined apoA-IV polymorphisms to platelet αIIbβ3 suggest a potential mechanism for increased risk of cardiovascular diseases in individuals with apoA-IVQ360H and apoA-IVT347S polymorphisms.
Keywords:
apolipoprotein A-IV; fibrinogen; fluorescence anisotropy; integrin αIIbβ3; molecular docking; molecular modeling; platelet; receptor structure–function; structural model; thrombosis.