Monitoring fibril formation of the N-terminal domain of PABPN1 carrying an alanine repeat by tryptophan fluorescence and real-time NMR.
Intranuclear fibrils due to poly-alanine expansions in the N-terminal domain of the poly(A) binding protein PABPN1 correlate with the disease oculopharyngeal muscular dystrophy (OPMD). For monitoring fibril formation by fluorescence and real-time NMR spectroscopy, tryptophans were introduced either into the middle or C-terminal of the poly-alanine segment. The kinetics of fibril formation which were monitored by fluorescence spectroscopy were matched by real-time NMR kinetics. Our results show that fibril formation is concomitant with the burial of the tryptophans in the fibrillar core. Since no soluble pre-fibrillar intermediate(s) was detected, fibril formation of this domain may be regarded as a two state conversion from an unfolded soluble into folded insoluble species.
Rohrberg
Julia
J
Institute for Biochemistry and Biotechnology of the Martin-Luther-University Halle-Wittenberg, Kurt-Mothes-Strasse 3, 06120 Halle, Germany.
Sachs
Rolf
R
Lodderstedt
Grit
G
Sackewitz
Mirko
M
Balbach
Jochen
J
Schwarz
Elisabeth
E
D000409:Alanine / Q000737:chemistry / Q000235:genetics; D000595:Amino Acid Sequence; D000682:Amyloid / Q000737:chemistry / Q000378:metabolism*; D005453:Fluorescence; D006801:Humans; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods*; D039103:Poly(A)-Binding Protein II / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D020449:Repetitive Sequences, Amino Acid; D014364:Tryptophan / Q000032:analysis*
D000682:Amyloid; D039103:Poly(A)-Binding Protein II; D011994:Recombinant Proteins; D014364:Tryptophan; D000409:Alanine