Clostridium cellulovorans produces multi-enzyme complexes called cellulosomes capable of efficiently degrading cellulosic biomass. There are three xylanase genes containing a sequence corresponding to a dockerin domain that are necessary for constructing cellulosomes in the genome. Among the xylanases encoded by these genes, xylanase B (XynB) contains a catalytic domain belonging to glycoside hydrolase family 10 and a carbohydrate-binding module (CBM) at the N-terminus, making it a member of CBM family 22. In this study, XynB was cloned, overexpressed, purified and crystallized. XynB was crystallized using the hanging-drop vapour-diffusion method in the presence of 0.2 M sodium acetate trihydrate, 0.1 M Tris-HCl pH 8.5, 32%(w/v) PEG 4000 at 293 K. X-ray diffraction analysis revealed that the crystal diffracted to 1.95 Å resolution and belonged to space group P222, with unit-cell parameters a = 74.28, b = 77.55, c = 88.20 Å, α = β = γ = 90°. The data-evaluation statistics revealed high quality of the collected data, thereby establishing a solid basis for determination of the structure of cellulosomal xylanase from C. cellulovorans.
SUMMARY
In the biological and medical fields, sterilization using pulsed electric fields or discharges and electroporation have been studied actively in recent years. It is clear that a pulsed electric field has some sort of effect on an organism. At present, however, it is not sufficiently clear what effect occurs and how the mechanism of a pulsed electric field acts on the organism. The effects of pulsed electric fields on ribonucleic acid (RNA) expression in biological cells were studied with the objective of elucidating the bioelectric mechanism. A pulsed electric field was applied to Vibrio parahaemolyticus, a pathogenic bacteria, and the expression of RNAs, specifically thermostable direct hemolysin (TDH), VP1680, and VP1699, which represent pathogenic agents, were analyzed using the polymerase chain reaction (PCR) method. The level of expression of VP1680 increased when a pulsed electric field was applied, and the effect was at a peak when the electric field strength was 14 kV/cm. The electric field strength barely affected the expression of TDH and VP1699. It was found that the effects of the electric field were different among the RNAs. The expression level varied with time when expression was analyzed. These results suggest the possibility of selective control of gene expression by using pulsed electric fields and possible new applications such as the development of functional foods.
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