Amyloid-forming property of the N-terminal 1−70 residues of human apolipoprotein A-IV

Apolipoprotein A-IV (apoA-IV), the largest member of the exchangeable apolipoprotein family, is a common constituent of amyloid deposits in renal and cardiac amyloidosis. In this study, we characterized the aggregation propensity of the apoA-IV N-terminal fragment to form amyloid fibrils using a variety of biophysical techniques. Thioflavin T fluorescence assay, circular dichroism measurement, and microscopic observations revealed that the N-terminal 1−70 amino acid fragment of apoA-IV readily forms amyloid fibrils by a transition from a random coil to a β-sheet-rich structure. Sequence-based analysis indicated that residues 7−16 and 38−42 are the major aggregation-prone segments within the N-terminal 1−70 residues of apoA-IV. Consistent with this, deletion of these residues strongly inhibited the β-transition and fibril formation of apoA-IV 1−70. Kinetic and thermodynamic analyses of fibril formation by the apoA-IV 1−70 fragment demonstrated that primary nucleation is the dominant step in fibril formation, for which the activation energy barrier is entirely entropic. In addition, we found that the presence of heparin, a representative glycosaminoglycan, accelerated fibril formation kinetics and enhanced the yield of apoA-IV 1−70 fibrils, and the positively charged residues K58-K59 play a critical role in heparin interaction. Overall, our results suggest that the strong amyloid-forming propensity of the N-terminal fragment of apoA-IV may play a key role in amyloid deposition associated with apoA-IV amyloidosis.