Takashi Sakamoto

In this Feature Article, we focus on recent advances in our research on molecular recognition and fluorescence sensing of phosphate anion derivatives of biological importance. Because of their significant roles in biological systems, considerable efforts have been devoted to developing detection or determination systems. However, the recognition and sensing of these anion species under aqueous biological conditions using small-molecular chemosensors still remain as a challenging research topic. We have been developing a variety of artificial receptors and fluorescent chemosensors for phosphoproteins and nucleoside polyphosphates in recent years. They consist of a binuclear Zn(II)-dipicolylamine (Dpa) complex as a common binding motif for phosphate anion derivatives. Taking advantage of their strong binding affinities or high sensing abilities, a variety of biological assay systems have also been successfully developed, which includes the enzyme assays such as the kinase, phosphatase and glycosyltransferase reaction, as well as an inhibitor assay for the phosphoprotein-protein surface interaction.

We have developed a new fluorescent binuclear Zn(II) complex for the detection of neurofibrillary tangles (NFTs) of hyperphosphorylated tau proteins, a representative hallmark of Alzheimer's disease (AD). The probe 1 incorporates a fluorescent BODIPY unit and two Zn(II)-2,2'-dipicolylamine (Dpa) complexes as a binding site for phosphorylated amino acid residues. Using fluorescence titration to evaluate the binding and sensing properties of 1 toward several phosphorylated peptide segments derived from hyperphosphorylated tau protein, we found that 1 binds preferentially to peptides presenting phosphorylated groups at the i and i+4 positions with dissociation constants (K(d)) in the micromolar range. Fluorescence titration with an artificially prepared aggregate of the phosphorylated tau protein (p-Tau) revealed that 1 binds strongly to p-Tau (EC(50) = 9 nM). In contrast, the interactions of 1 were weaker toward artificially prepared aggregates of the nonphosphorylated tau protein (n-Tau; EC(50) = 80 nM) and Abeta(1-42) fibrils (EC(50) = 650 nM). Histological imaging of a hippocampus tissue section obtained from an AD patient revealed that 1 fluorescently visualizes deposits of NFTs and clearly discriminates between NFTs and the amyloid plaques assembled from amyloid-beta peptides, confirming our strategy toward the rational design of a molecular probe for the selective fluorescence detection of NFTs in brain tissue sections.

Highly syndiotactic poly(methyl methacrylate)s (PMMAs) with narrow molecular weight distribution (MWD) were prepared by polymerization of MMA in toluene at low temperatures, initiated with tert-butyllithium (t-C4H9Li) combined with trialkylaluminium compounds such as triethyl-, tributyl-, and trioctylaluminium (mole ratio Al/Li ≥ 3). Polymerization was initiated by the t-C4H9 anion and proceeded in a living manner. Several alkyl methacrylates were also polymerized with t-C4H9Li-(n-C4H9)3Al to give highly syndiotactic polymethacrylates with narrow MWD. Highly syndiotactic block and random copolymers of MMA and ethyl methacrylate were obtained by this initiator.

A cell growth inhibitory effect of drupanin and baccharin, ingredients of propolis, was found in human cancer cell lines. These compounds induced apoptosis in the cells characterized by morphological and nucleosomal DNA fragmentation analysis. Their effects were less potent compared with that of artepillin C, which is a known anticancer compound from propolis. Importantly, HL60 cells were more sensitive to drupanin than were Con A-stimulated peripheral blood lymphocytes, whereas the potency of artepillin C was the opposite of that of drupanin.