Structural Basis for the Pro-amyloidogenic Action and Ligand Binding of a Novel W72R Variant of Human Apolipoprotein A-I.

Apolipoprotein A-I (apoA-I), the major structural and functional protein of plasma high-density lipoprotein (HDL), mediates cholesterol removal from the body and has other beneficial properties. APOA1 gene mutations can cause diseases including human hereditary AApoAI amyloidosis, a life-threatening multiorgan disorder caused by apoA-I misfolding and amyloid deposition. We report esophageal AApoAI amyloidosis in a 66-year-old patient. Amyloid fibrils are comprised of a novel p.Trp96Arg (W72R) apoA-I variant identified by protein mass spectrometry and gene sequencing. Both variant and wild-type apoA-I circulate in plasma at low HDL levels typical of this dominant negative disorder. To understand why W72R substitution promotes amyloid formation, we generated recombinant full-length W72R and wild-type proteins and compared their structural, stability and ligand-binding properties using spectroscopic (circular dichroism, fluorescence, absorption) and biochemical approaches. High-resolution structures of apoA-I in native and in amyloid states were harnessed to propose structural underpinning for the pro-amyloidogenic effects of W72R substitution. W72R substitution slightly destabilized native lipid-free protein; decreased structural protection of amyloid-promoting regions; accelerated lipid solubilization by apoA-I and destabilized model lipoproteins; facilitated apoA-I binding to heparin (a proxy for heparan sulfate) and to type-I collagen; accelerated amyloid formation and potentially stabilized the amyloid structure. All these effects are pro-amyloidogenic. The results help explain the pro-amyloidogenic nature of this mutation, suggest apoA-I binding to extracellular matrix components such as heparan sulfate and collagen as therapeutic targets, and expand our knowledge of the clinical presentation and molecular basis of this complex disease.