The effect of bone matrix protein of osteonectin on de novo formation of apatite was studied in a wide range of calcium phosphate solutions in the presence of collagen. In every solution, from which amorphous calcium phosphate, octacalcium phosphate, or apatite precipitated as a possible initial phase, osteonectin at concentrations less than 1 microM retarded the precipitation, subsequent transformation to apatite, and ripening crystal growth of apatite. Collagen present as either reconstituted or denatured form had no effect on the osteonectin-associated reactions as well as osteonectin-free reactions, and no structural correlation was observed between collagen fibrils and any of the calcium phosphates that appeared in our system. Direct measurement of free calcium levels in the solutions suggested that the reduction in calcium activity due to complexing with osteonectin hardly explained the inhibitory activity of osteonectin in retarding the formation of apatite. Instead, our transmission electron microscopic (TEM) observation strongly suggested that the primary mechanism for osteonectin to inhibit the formation of apatite is to block growth sites of calcium phosphates nucleated. The apatite thus formed in the presence of osteonectin showed less resolved X-ray diffraction patterns, partly because of smaller crystallites as suggested by TEM.

a b s t r a c tInvestigation of the mechanism of tau polymerization is indispensable for finding inhibitory conditions or identifying compounds preventing the formation of paired helical filament or oligomers. Tau contains a microtubule-binding domain consisting of three or four repeats in its C-terminal half. It has been considered that the key event in tau polymerization is the formation of a b-sheet structure arising from a short hexapeptide 306 VQIVYK311 in the third repeat of tau. In this paper, we report for the first time that the C-HÁ Á Áp interaction between Ile308 and Tyr310 is the elemental structural scaffold essential for forming a dry ''steric zipper" structure in tau amyloid fibrils.

Information on the structural scaffold for tau aggregation is important in developing a method of preventing Alzheimer's disease (AD). Tau contains a microtubule binding domain (MBD) consisting of three or four repeats of 31 and 32 similar residues in its C-terminal half. Although the key event in tau aggregation has been considered to be the formation of β-sheet structures from a short hexapeptide (306)VQIVYK(311) in the third repeat of MBD, its aggregation pathway to filament formation differs between the three- and four-repeated MBDs, owing to the intermolecular and intramolecular disulphide bond formations, respectively. Therefore, the elucidation of a common structural element necessary for the self-assembly of three-/four-repeated full-length tau is an important research subject. Expanding the previous results on the aggregation mechanism of MBD, in this paper, we report that the C-H … π interaction between the Ile308 and Tyr310 side chains in the third repeat of MBD is indispensable for the self-assembly of three-/four-repeated full-length tau, where the interaction provides a conformational seed for triggering the molecular association. On the basis of the aggregation behaviours of a series of MBD and full-length tau mutants, a possible self-association model of tau is proposed and the relationship between the aggregation form (filament or granule) and the association pathway is discussed.