In drug discovery, efficient screening of protein-drug interactions (PDIs) is hampered by the limitations of current biophysical approaches. Here, we develop a biological nanopore sensor for single-molecule detection of proteins and PDIs using the pore-forming toxin YaxAB. Using this YaxAB nanopore, we demonstrate label-free, single-molecule detection of interactions between the anticancer Bcl-xL protein and small-molecule drugs as well as the Bak-BH3 peptide. The long funnel-shaped structure and nanofluidic characteristics of the YaxAB nanopore enable the electro-osmotic trapping of diverse folded proteins and high-resolution monitoring of PDIs. Distinctive nanopore event distributions observed in the two-dimensional (ΔI/Io-versus-IN) plot illustrate the ability of the YaxAB nanopore to discriminate individual small-molecule drugs bound to Bcl-xL from non-binders. Taken together, our results present the YaxAB nanopore as a robust platform for label-free, ultrasensitive, single-molecule detection of PDIs, opening up a possibility for low-cost, highly efficient drug discovery against diverse drug targets.

ObjectiveThe purpose of this study was to evaluate the effects of three different surface conditioning methods on the shear bond strength (SBS) of metal brackets bonded directly to gold alloy with chemically cured resin.MethodsTwo hundred ten type III gold alloy specimens were randomly divided into six groups according to the combination of three different surface conditioning methods (aluminum oxide sandblasting only, application of a metal primer after aluminum oxide sandblasting, silica coating and silanation) and thermocycling (with thermocycling, without thermocycling). After performing surface conditioning of specimens in accordance with each experimental condition, metal brackets were bonded to all specimens using a chemically cured resin. The SBS was measured at the moment of bracket debonding, and the resin remnants on the specimen surface were evaluated using the adhesive remnant index.ResultsApplication of metal primer after aluminum oxide sandblasting yielded a higher bond strength than that with aluminum oxide sandblasting alone (p < 0.001), and silica coating and silanation yielded a higher bond strength than that with metal primer after aluminum oxide sandblasting (p < 0.001). There was no significant change in SBS after thermocycling in all groups.ConclusionsWith silica coating and silanation, clinically satisfactory bond strength can be attained when metal brackets are directly bonded to gold alloys using a chemically cured resin.

Hydrolysis and absorption properties of submicron-(below 1 lm) and micron-sized (over 100 lm) powders of raw rice, heat-gelatinized rice, fermented rice and wheat were investigated in vitro and in vivo. Nutrient content such as protein, carbohydrate or vitamin C in the samples was not different significantly between submicron-and micron-sized powders, but it showed difference according to processed type of the powders. Submicron-sized powders were hydrolyzed more easily than micron-sized powders by a-amylases such as industrial, human salivary and porcine pancreatic a-amylase but not by a-glucosidase in all the tested samples. Patterns of blood glucose change were investigated in rats. The highest glucose levels were obtained at 30 min after oral administration in both submicron-and micron-sized powders, but the levels were decreased drastically in groups administered submicron-sized powders, while they were slow and gradual in groups administered micron-sized powders. On the contrary, the body weight gained and food efficiency ratios showed a tendency to be decreased slightly in mice fed with submicron-sized rice powders compared to those with micron-sized rice powder.

The RING domain of MUL1 (RING MUL1 ) alone mediates ubiquitylation of the p53-transactivation domain (TAD p53 ). To elucidate the mechanism underlying the simultaneous recruitment of UBE2D2 and the substrate TAD p53 by RING MUL1 , we determined the complex structure of RING MUL1 :UBE2D2 and studied the interaction between RING MUL1 and TAD p53 in the presence of UBE2D2-UB thioester (UBE2D2~UB) mimetics. The RING MUL1 -binding induced the closed conformation of UBE2D2 S22R/C85S -UB K48R oxyester (UBE2D2 RS -UB R OE ), and strongly accelerated its hydrolysis, which was suppressed by the additional N77Amutation of UBE2D2. Interestingly, UBE2D2 S22R/N77A/C85S -UB K48R oxyester (UBE2D2 RAS -UB R OE ) already formed a closed conformation in the absence of RING MUL1 . Although TAD p53 exhibited weak binding for RING MUL1 or UBE2D2 alone, its binding affinity was enhanced and even further for RING MUL1 :UBE2D2 and RING MUL1 :UBE2D2 RAS -UB R OE , respectively. The recognition of TAD p53 by RING MUL1 as a complex with UBE2D2~UB is related to the multivalency of the binding events and underlies the ability of RING MUL1 to ubiquitylate the intrinsically disordered protein, TAD p53 .