An extracellular chitinase produced by Paenibacillus pasadenensis CS0611was purified by ammoniumsulfate precipitation,HiTrap DEAE FF and HiLoad26/600Superdex200pg column chromatography.The apparent molecular mass dete...An extracellular chitinase produced by Paenibacillus pasadenensis CS0611was purified by ammoniumsulfate precipitation,HiTrap DEAE FF and HiLoad26/600Superdex200pg column chromatography.The apparent molecular mass determined by sodium dodecyl sulfate polyacrylamide gelelectrophoresis was69kDa.The optimum pH and optimum temperature of the chitinase were5.0and50°C,respectively.The enzyme showed high stability at alkaline pH values and temperaturesbelow40°C.Additionally,the metal ions Mn2+,Mg2+,and Co2+inhibited activity of the chitinase.Thechitinase was active on colloidal chitin with an apparent Km of4.41mg/mL and Vmax of1.08mg/min.Substrate spectrum analysis indicated that the chitinase reacted preferentially with the glucosidicbond between GlcNAc‐GlcNAc.The enzymatic hydrolysate was analyzed by high‐performance liquidchromatography and thin layer chromatography,and clearly showed that a subunit of(GlcNAc)2was the main hydrolysis product.展开更多
We found a novel lipase gene in the Paenibacillus pasadenensis CS0611 strain.The lipase gene sequence was cloned into the pET-28a expression vector to construct a recombinant lipase protein containing 6×His tags ...We found a novel lipase gene in the Paenibacillus pasadenensis CS0611 strain.The lipase gene sequence was cloned into the pET-28a expression vector to construct a recombinant lipase protein containing 6×His tags at the C-and N-termini,respectively.High-level expression of the lipase in E.coli BL21(DE3)was obtained upon induction with IPTG at 20°C.The recombinant lipase activity was approximately 1631-fold higher than the wild type.His-tagged recombinant lipase was purified rapidly and efficiently by using Ni-charged affinity chromatography with 63.5%recovery and a purification factor of 10.78.The purified lipase was stable in a broad range of temperatures and pH values,with the optimal temperature and pH being 50°C and 7.0,respectively.Its activity was stimulated to different degrees in the presence of metal ions such as Ca2+,Mg2+,and some non-ionic surfactants.In addition,the purified lipase was activated by a series of water-miscible organic solvents such as some short carbon chain alcohols and was highly tolerant to some water-immiscible organic solvents.展开更多
Fifty bacterial strains isolated from dairy product, skin and blood from cancer and kidney failure dialysis patients were identified to 22 species and the following genera： Brevibacterium, Corynebacterium, Arthrobact...Fifty bacterial strains isolated from dairy product, skin and blood from cancer and kidney failure dialysis patients were identified to 22 species and the following genera： Brevibacterium, Corynebacterium, Arthrobacter, Actinomyces, Exiguobacterium, Kocuria, Micrococcus, Rothia, Rhodococcus and classified numerically using a set of 52 phenetic characteristics, using simple matching coefficient （Ssm） and clustering method of unweighted average linkage between groups by SPSS program. They were also grouped to 7 main clusters and 29 sub-clusters in the hierarchical tree. Twelve isolates of the different species from the genera Brevibacterium, Arthrobacter, Corynebacterium, Kocuria, Rhodococcus, Rothia were selected from the taxonomic clusters and probed for lin gene by peR. One species Kocuria rhizophila which inhibited most of the test organism did not have lin gene in the chromosome while the species Corynebacterium glucuronolyticum, Arthrobacter comminsii, Arthrobacter oxydans have the lin gene. Our results establish a wide distribution of the structural gene encoding this Iinocin M 18 within coryneform bacteria and also in the genus Kocuria.展开更多
基金supported by the National Natural Science Foundation of China (21336002,21376096,21676104)~~
文摘An extracellular chitinase produced by Paenibacillus pasadenensis CS0611was purified by ammoniumsulfate precipitation,HiTrap DEAE FF and HiLoad26/600Superdex200pg column chromatography.The apparent molecular mass determined by sodium dodecyl sulfate polyacrylamide gelelectrophoresis was69kDa.The optimum pH and optimum temperature of the chitinase were5.0and50°C,respectively.The enzyme showed high stability at alkaline pH values and temperaturesbelow40°C.Additionally,the metal ions Mn2+,Mg2+,and Co2+inhibited activity of the chitinase.Thechitinase was active on colloidal chitin with an apparent Km of4.41mg/mL and Vmax of1.08mg/min.Substrate spectrum analysis indicated that the chitinase reacted preferentially with the glucosidicbond between GlcNAc‐GlcNAc.The enzymatic hydrolysate was analyzed by high‐performance liquidchromatography and thin layer chromatography,and clearly showed that a subunit of(GlcNAc)2was the main hydrolysis product.
文摘We found a novel lipase gene in the Paenibacillus pasadenensis CS0611 strain.The lipase gene sequence was cloned into the pET-28a expression vector to construct a recombinant lipase protein containing 6×His tags at the C-and N-termini,respectively.High-level expression of the lipase in E.coli BL21(DE3)was obtained upon induction with IPTG at 20°C.The recombinant lipase activity was approximately 1631-fold higher than the wild type.His-tagged recombinant lipase was purified rapidly and efficiently by using Ni-charged affinity chromatography with 63.5%recovery and a purification factor of 10.78.The purified lipase was stable in a broad range of temperatures and pH values,with the optimal temperature and pH being 50°C and 7.0,respectively.Its activity was stimulated to different degrees in the presence of metal ions such as Ca2+,Mg2+,and some non-ionic surfactants.In addition,the purified lipase was activated by a series of water-miscible organic solvents such as some short carbon chain alcohols and was highly tolerant to some water-immiscible organic solvents.
文摘Fifty bacterial strains isolated from dairy product, skin and blood from cancer and kidney failure dialysis patients were identified to 22 species and the following genera： Brevibacterium, Corynebacterium, Arthrobacter, Actinomyces, Exiguobacterium, Kocuria, Micrococcus, Rothia, Rhodococcus and classified numerically using a set of 52 phenetic characteristics, using simple matching coefficient （Ssm） and clustering method of unweighted average linkage between groups by SPSS program. They were also grouped to 7 main clusters and 29 sub-clusters in the hierarchical tree. Twelve isolates of the different species from the genera Brevibacterium, Arthrobacter, Corynebacterium, Kocuria, Rhodococcus, Rothia were selected from the taxonomic clusters and probed for lin gene by peR. One species Kocuria rhizophila which inhibited most of the test organism did not have lin gene in the chromosome while the species Corynebacterium glucuronolyticum, Arthrobacter comminsii, Arthrobacter oxydans have the lin gene. Our results establish a wide distribution of the structural gene encoding this Iinocin M 18 within coryneform bacteria and also in the genus Kocuria.
