One-step synthesis of positively charged bifunctional carbon dot/silver composite nanoparticles for killing and fluorescence imaging of Gram-negative bacteria.

Positively charged C-dot/Ag composite nanoparticles were synthesized via the facile one-step hydrothermal reaction of L-arginine and silver nitrate. L-arginine was used not only as the carbon and nitrogen sources of N-doped C-dots but also as the reducing agent of silver ions. It was noteworthy that the resulting C-dots were negatively charged but the simultaneous reduction of silver ions made the resulting C-dot/Ag composite nanoparticles become positively charged. Furthermore, as compared to C-dots, the presence of Ag nanoparticles and the higher nitrogen content led to the redshift of excitation and emission intervals. Also, the enlarged excitation wavelength range in the visible light region made the resulting C-dot/Ag nanocomposite more useful in fluorescence imaging. In addition, the C-dot/Ag composite nanoparticles exhibited more excellent bacteria-killing capability than C-dots and were successfully used for the fluorescence imaging of E. coli because they could attach and release silver ions on the surface of E. coli. In conclusion, a facile one-step hydrothermal process has been successfully developed for the synthesis of C-dot/Ag composite nanoparticles, and the resulting C-dot/Ag composite nanoparticles are expected to have great potential in the killing and fluorescence imaging of Gram-negative bacteria.

Characterization of a recombinant d-allulose 3-epimerase from Agrobacterium sp. ATCC 31749 and identification of an important interfacial residue.

d-Allulose 3-epimerase (DAEase) catalyzes the epimerization between d-fructose and d-allulose. We had PCR-cloned and overexpressed the gene encoding Agrobacterium sp. ATCC 31749 DAEase (AsDAEase) in Escherichia coli. A high yield of active AsDAEase, 35,300U/L or 1350U/g of wet cells, was acquired with isopropyl ß-d-1-thiogalactopyranoside induction at 20°C for 20h. Although only six residues including residue 234 located in tetrameric interface are different between AsDAEase and A. tumefaciens DAEase (AtDAEase), the specific activity of purified AsDAEase is much larger than that of AtDAEase. The optimal pHs and optimal temperatures of the purified recombinant AsDAEase are 7.5-8.0 and 55-60°C, respectively. The half-life of the enzyme is 267min at 55°C in the presence of 0.1mM Co2+, and the equilibrium ratio between d-allulose and d-fructose is 30:70 at 55°C. Besides characterizing AsDAEase, mutation N234D was constructed to assess its influence on activity. The specific activity of the purified N234D AsDAEase is only 25.5% of wild-type's activity, suggesting residue N234 is an important interfacial residue which substantially affects enzyme activity. The high specific activity and high expression yield of AsDAEase suggest its prospect to be applied in d-allulose production.