A simple and rapid method (slow-stirring method) for extracting environmental DNA (eDNA) from soils was constructed by physical mild stirring with chemical treatment. eDNA was extracted efficiently with minimal damage from various kinds of soil. The amount of eDNA and soil bacterial biomass showed a linear proportional relation [Y=(1.70x10(8))X, r2=0.96], indicating that bacterial biomass could be evaluated by quantifying levels of eDNA. Consequently, the average bacterial biomass in an agricultural field was calculated as 5.95x10(9) cells/g sample, approximately 10-100 times higher than that in non- and oil-polluted fields.
Aims: To degrade ether‐type polyurethane (ether‐PUR), ether‐PUR–degrading micro‐organism was isolated. Moreover, ether‐PUR–degrading mechanisms were analysed using model compounds of ether‐PUR.
Methods and Results: A fungus designated as strain PURDK2, capable of changing the configuration of ether‐PUR, has been isolated. This isolated fungus was identified as Alternaria sp. Using a scanning electron microscope, the grid structure of ether‐PUR was shown to be melted and disrupted by the fungus. The degradation of ether‐PUR by the fungus was analysed, and the ether‐PUR was degraded by the fungus by about 27·5%. To analyse the urethane‐bond degradation by the fungus, a degraded product of ethylphenylcarbamate was analysed using GC/MS. Aniline and ethanol were detected by degradation with the supernatant, indicating that the fungus secreted urethane‐bond–degrading enzyme(s). PURDK2 also degraded urea bonds when diphenylmethane‐4,4′‐dibutylurea was used as a substrate.
Conclusions: The enzyme(s) from PURDK2 degraded urethane and urea bonds to convert the high molecular weight structure of ether‐PUR to small molecules; and then the fungus seems to use the small molecules as an energy source.
Significance and Impact of the Study: Ether‐PUR–degrading fungus, strain PURDK2, was isolated, and the urethane‐ and urea‐bonds–degrading enzymes from strain PURDK2 could contribute to the material recycling of ether‐PUR.
Thirty-six bacteria that degraded long-chain hydrocarbons were isolated from natural environments using long-chain hydrocarbons (waste car engine oil, base oil or the c-alkane fraction of base oil) as the sole carbon and energy source. A phylogenetic tree of the isolates constructed using their 16S rDNA sequences revealed that the isolates were divided into six genera plus one family (Acinetobacter, Rhodococcus, Gordonia, Pseudomonas, Ralstonia, Bacillus and Alcaligenaceae, respectively). Furthermore, most of the isolates (27 of 36) were classified into the genera Acinetobacter, Rhodococcus or Gordonia. The hydrocarbon-degradation similarity in each strain was confirmed by the 2,6-dichlorophenol indophenol (2,6-DCPIP) assay. Isolates belonging to the genus Acinetobacter degraded long-chain normal alkanes (n-alkanes) but did not degrade short-chain n-alkanes or cyclic alkanes (c-alkanes), while isolates belonging to the genera Rhodococcus and Gordonia degraded both long-chain n-alkanes and c-alkanes.
The relationship between the autodegradation and thermostability of thermolysin (TLN) was studied. Four autodegradation sites in TLN were identified in the presence of Ca(2+). One of the sites was identified as Gly(154)-Leu(155), and Leu(155) was substituted with various amino acids, X = Ala, Ser, Phe, and Gly, by site-directed mutagenesis. The thermostability at 80 degrees C increased with the amino acid substitutions in the order of Ala>Phe>Ser>Gly>Leu (WT TLN). An additional autodegradation fragment that was not observed with WT TLN appeared for all mutant TLNs examined. The autodegradation site shifted from the Gly(154)-Leu(155) bond to the X(155)-Ile(156) one with the mutation at Leu(155). Furthermore, the Ile(164)-Asp(165) bond was recognized newly as an autodegradation site in the mutant TLNs for the production of AF3'.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.