Quantitative characterization of heparin binding to Tau protein: implication for inducer-mediated Tau filament formation.
Neurofibrillary tangles, principally composed of bundles of filaments formed by the microtubule-associated protein Tau, are a hallmark of a group of neurodegenerative diseases such as Alzheimer disease. Polyanionic cofactors such as heparin can induce Tau filament formation in vitro. Here we quantitatively characterize the interaction between recombinant human Tau fragment Tau(244-372) and heparin (average molecular mass = 7 kDa) as well as heparin-induced fibril formation by using static light scattering, isothermal titration calorimetry, turbidity assays, and transmission electron microscopy. Our data clearly show that at physiological pH, heparin 7K, and human Tau(244-372) form a tight 1:1 complex with an equilibrium association constant exceeding 10(6) m(-1) under reducing conditions, triggering Tau fibrillization. In the absence of dithiothreitol, heparin shows a moderate binding affinity (10(5) m(-1)) to Tau(244-372), similarly triggering Tau fibrillization. Further fibrillization kinetics analyses show that the lag time appears to be approximately invariant up to a molar ratio of 2:1 and then increases at larger ratios of heparin/Tau. The maximum slope representing the apparent rate constant for fibril growth increases sharply with substoichiometric ratios of heparin/Tau and then decreases to some extent with ratios of >1:1. The retarding effect of heparin in excess is attributed to the large increase in ionic strength of the medium arising from free heparin. Together, these results suggest that the formation of the 1:1 complex of Tau monomer and heparin plays an important role in the inducer-mediated Tau filament formation, providing clues to understanding the pathogenesis of neurodegenerative diseases.
Zhu
Hai-Li
HL
From the State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan 430072, China and.
Fernández
Cristina
C
the Laboratory of Biochemistry and Genetics, NIDDK, National Institutes of Health, Bethesda, Maryland 20892.
Fan
Jun-Bao
JB
From the State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan 430072, China and.
Shewmaker
Frank
F
the Laboratory of Biochemistry and Genetics, NIDDK, National Institutes of Health, Bethesda, Maryland 20892.
Chen
Jie
J
From the State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan 430072, China and.
Minton
Allen P
AP
the Laboratory of Biochemistry and Genetics, NIDDK, National Institutes of Health, Bethesda, Maryland 20892.
Liang
Yi
Y
From the State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan 430072, China and. Electronic address: liangyi@whu.edu.cn.
D000465:Algorithms; D002151:Calorimetry / Q000379:methods; D006493:Heparin / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D046529:Microscopy, Electron, Transmission; D008956:Models, Chemical; D008970:Molecular Weight; D016874:Neurofibrillary Tangles / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure; D010084:Oxidation-Reduction; D010446:Peptide Fragments / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D016875:tau Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
D010446:Peptide Fragments; D011994:Recombinant Proteins; D016875:tau Proteins; D006493:Heparin