Single bacterial resolvases first exploit, then constrain intrinsic dynamics of the Holliday junction to direct recombination

Homologous recombination forms and resolves an entangled DNA Holliday Junction (HJ) crucial for achieving genetic reshuffling and genome repair. To maintain genomic integrity, specialized resolvase enzymes cleave the entangled DNA into two discrete DNA molecules. However, it is unclear how two similar stacking isomers are distinguished, and how a cognate sequence is found and recognized to achieve accurate recombination. We here use single-molecule fluorescence observation and cluster analysis to examine how prototypic bacterial resolvase RuvC singles out two of the four HJ strands and achieves sequence-specific cleavage. We find that RuvC first exploits, then constrains the dynamics of intrinsic HJ isomer exchange at a sampled branch position to direct cleavage toward the catalytically competent HJ conformation and sequence, thus controlling recombination output at minimal energetic cost. Our model of rapid DNA scanning followed by ‘snap-locking’ of a cognate sequence is strikingly consistent with the conformational proofreading of other DNA-modifying enzymes.

Study on the dependence of fluorescence intensity on optical density of solutions: the use of fluorescence observation field for inner filter effect corrections

Absorbance value that corresponds to maximum fluorescence intensity (Aflmax) depends on fluorescence observation field.

Fabrication of Core-Shell PLGA-Chitosan Microparticles Using Electrospinning: Effects of Polymer Concentration

This investigation aims to fabricate the core-shell microparticles composed of poly(lactic-co-glycolic acid) and chitosan (PLGA-CS MPs) using electrospinning. The challenge of using electrospinning is that it has many parameters which change product outcome if any single parameter is changed. However, the advantage of this system is that we can fabricate either micro/nanofibers or micro/nanoparticles. To learn about the effect of liquid concentration, the electrospinning parameters (voltage, needle sizes, distance from needle to collector, and ejection speed) were fixed while the concentration of PLGA or chitosan was varied. The results showed that PLGA microparticles can be fabricated successfully when the concentration of PLGA is smaller than 10 wt%. Presence of the chitosan shell was confirmed by zeta potential measurements, FT-IR, optical observation, and fluorescence observation. Thickness of the chitosan shell can be controlled by changing the concentration of chitosan and measured by fluorescamine labeling method. Moreover, SEM observation showed that concentration of chitosan affected the size of PLGA-CS microparticles. The MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) assay test showed that PLGA-CS microparticles possess excellent biocompatibility.