Identification of beta-amyloid-binding sites on transthyretin.
Transthyretin (TTR) binds to the Alzheimer-related peptide beta-amyloid (Aβ), and may protect against Aβ-induced neurotoxicity. In this work, the specific domains on TTR involved with binding to Aβ were probed. An array was constructed of peptides derived from overlapping sequences from TTR. Strong binding of Aβ to TIAALLSPYSYS (residues 106-117) was detected, corresponding to strand G on the inner β-sheet of TTR. Aβ bound weakly to four contiguous peptides spanning residues 59-83, which includes strand E through the E/F helix and loop. To further pinpoint specific residues on TTR involved with Aβ binding, nine alanine mutants were generated: I68A, I73A, K76A, L82A, I84A, S85A, L17A, T106A and L110A. Aβ binding was significantly inhibited only in L82A and L110A, indicating that Aβ binding to TTR is mediated through these bulky hydrophobic leucines. Aβ binding to L17A and S85A was significantly higher than to wild-type TTR. Enhancement of binding in L17A is postulated to arise from reduced steric restriction to the interior L110 site, since these two residues are adjacent in the native protein. The S85A mutation caused a reduction in TTR tetramer stability; increased Aβ binding is postulated to be a direct consequence of the reduced quaternary stability.
Chemical and Biological Engineering Department, University of Wisconsin-Madison, Madison, WI 53706, USA.
Jiali
J
Du
Patricia Y
PY
Cho
Dennis T
DT
Yang
Regina M
RM
Murphy
D000409:Alanine / Q000235:genetics; D000595:Amino Acid Sequence; D016229:Amyloid beta-Peptides / Q000378:metabolism*; D001665:Binding Sites; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D011228:Prealbumin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary
D016229:Amyloid beta-Peptides; D011228:Prealbumin; D000409:Alanine