Alzheimer’s disease (AD) is the most prevalent neurodegenerative disease in the elderly. Zinc (Zn) ion interacts with the pathogenic hallmark, amyloid-β (Aβ), and is enriched in senile plaques in brain of AD patients. To understand Zn-chelated Aβ (ZnAβ) species, here we systematically characterized ZnAβ aggregates by incubating equimolar Aβ with Zn. We found ZnAβ40 and ZnAβ42 both form spherical oligomers with a diameter of ~12–14 nm composed of reduced β-sheet content. Oligomer assembly examined by analytical ultracentrifugation, hydrophobic exposure by BisANS spectra, and immunoreactivity of ZnAβ and Aβ derived diffusible ligands (ADDLs) are distinct. The site-specific 13C labeled solid-state NMR spectra showed that ZnAβ40 adopts β-sheet structure as in Aβ40 fibrils. Interestingly, removal of Zn by EDTA rapidly shifted the equilibrium back to fibrillization pathway with a faster kinetics. Moreover, ZnAβ oligomers have stronger toxicity than ADDLs by cell viability and cytotoxicity assays. The ex vivo study showed that ZnAβ oligomers potently inhibited hippocampal LTP in the wild-type C57BL/6JNarl mice. Finally, we demonstrated that ZnAβ oligomers stimulate hippocampal microglia activation in an acute Aβ-injected model. Overall, our study demonstrates that ZnAβ rapidly form toxic and distinct off-pathway oligomers. The finding provides a potential target for AD therapeutic development.

Using Langevin dynamics simulations, we investigate the dynamics of chaperone-assisted translocation of a flexible polymer through a nanopore. We find that increasing the binding energy ε between the chaperone and the chain and the chaperone concentration N(c) can greatly improve the translocation probability. Particularly, with increasing the chaperone concentration a maximum translocation probability is observed for weak binding. For a fixed chaperone concentration, the histogram of translocation time τ has a transition from a long-tailed distribution to a gaussian distribution with increasing ε. τ rapidly decreases and then almost saturates with increasing binding energy for a short chain; however, it has a minimum for longer chains at a lower chaperone concentration. We also show that τ has a minimum as a function of the chaperone concentration. For different ε, a nonuniversal dependence of τ on the chain length N is also observed. These results can be interpreted by characteristic entropic effects for flexible polymers induced by either the crowding effect from a high chaperone concentration or the intersegmental binding for the high binding energy.

Background: Streptococcus pneumoniae BglA-2 is a GH-1 6-phospho-␤-glucosidase with specificity toward 1,4-linked 6-phospho-␤-glucosides. Results: BglA-2 and other GH-1 members adopt a similar overall structure and catalytic mechanism. Conclusion: Tyr 126 , Tyr 303 , and Trp 338 determine substrate specificity, and Ser 424 , Lys 430 , and Tyr 432 discriminate phosphorylated from non-phosphorylated substrate. A tryptophan residue discriminates 6-phospho-␤-glucosidase from 6-phospho-␤-galactosidase activities. Significance: BglA-2 structures provide new insight into characteristics and substrate specificity of 6-phospho-␤-glucosidase.

The ␤-N-acetylhexosaminidase (EC 3.2.1.52) from glycoside hydrolase family 20 (GH20) catalyzes the hydrolysis of the ␤-N-acetylglucosamine (NAG) group from the nonreducing end of various glycoconjugates. The putative surface-exposed N-acetylhexosaminidase StrH/Spr0057 from Streptococcus pneumoniae R6 was proved to contribute to the virulence by removal of ␤(1,2)-linked NAG on host defense molecules following the cleavage of sialic acid and galactose by neuraminidase and ␤-galactosidase, respectively. StrH is the only reported GH20 enzyme that contains a tandem repeat of two 53% sequence-identical catalytic domains (designated as GH20-1 and GH20-2, respectively). Here, we present the 2.1 Å crystal structure of the N-terminal domain of StrH (residues Glu-175 to Lys-642) complexed with NAG. It adopts an overall structure similar to other GH20 enzymes: a (␤/␣) 8 TIM barrel with the active site residing at the center of the ␤-barrel convex side. The kinetic investigation using 4-nitrophenyl N-acetyl-␤-D-glucosaminide as the substrate demonstrated that GH20-1 had an enzymatic activity (k cat /K m ) of one-fourth compared with GH20-2. The lower activity of GH20-1 could be attributed to the substitution of active site Cys-469 of GH20-1 to the counterpart Tyr-903 of GH20-2. A complex model of NAG␤(1,2)Man at the active site of GH20-1 combined with activity assays of the corresponding site-directed mutants characterized two key residues Trp-443 and Tyr-482 at subsite ؉1 of GH20-1 (Trp-876 and Tyr-914 of GH20-2) that might determine the ␤(1,2) substrate specificity. Taken together, these findings shed light on the mechanism of catalytic specificity toward the ␤(1,2)-linked ␤-N-acetylglucosides.