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Molecular and Biochemical Characterization of a Novel Intracellular Low-Temperature-Active Xylanase
( Jun Pei Zhou,),( Yan Yan Dong ),( Xiang Hua Tang ),( Jun Jun Li ),( Bo Xu ),( Qian Wu ),( Ya Jie Gao ),( Lu Pan ),( Zun Xi Huang ) 한국미생물 · 생명공학회 2012 Journal of microbiology and biotechnology Vol.22 No.4
A 990 bp full-length gene (xynAHJ2) encoding a 329- residue polypeptide (XynAHJ2) with a calculated mass of 38.4 kDa was cloned from Bacillus sp. HJ2 harbored in a saline soil. XynAHJ2 showed no signal peptide, distinct amino acid stretches of glycoside hydrolase (GH) family 10 intracellular endoxylanases, and the highest amino acid sequence identity of 65.3% with the identified GH 10 intracellular mesophilic endoxylanase iM-KRICT PX1-Ps from Paenibacillus sp. HPL-001 (ACJ06666). The recombinant enzyme (rXynAHJ2) was expressed in Escherichia coli and displayed the typical characteristics of low-temperatureactive enzyme (exhibiting optimum activity at 35 o C, 62% at 20 o C, and 38% at 10 o C; thermolability at ≥45 o C). Compared with the reported GH 10 low-temperature-active endoxylanases, which are all extracellular, rXynAHJ2 showed low amino acid sequence identities (<45%), low homology (different phylogenetic cluster), and difference of structure (decreased amount of total accessible surface area and exposed nonpolar accessible surface area). Compared with the reported GH 10 intracellular endoxylanases, which are all mesophilic and thermophilic, rXynAHJ2 has decreased numbers of arginine residues and salt bridges, and showed resistance to Ni 2+ , Ca 2+ , or EDTA at 10 mM final concentration. The above mechanism of structural adaptation for low-temperature activity of intracellular endoxylanase rXynAHJ2 is different from that of GH 10 extracellular low-temperature-active endoxylanases. This is the first report of the molecular and biochemical characterizations of a novel intracellular low-temperatureactive xylanase.
( Jun Mei Ding ),( Ting Ting Yu ),( Lian Ming Liang ),( Zhen Rong Xie ),( Yun Juan Yang ),( Jun Pei Zhou ),( Bo Xu ),( Jun Jun Li ),( Zun Xi Huang ) 한국미생물 · 생명공학회 2014 Journal of microbiology and biotechnology Vol.24 No.11
The esterase gene Est8 from the thermophilic bacterium Bacillus sp. K91 was cloned and expressed in Escherichia coli. The monomeric enzyme exhibited a theoretical molecular mass of 24.5 kDa and an optimal activity around 50°C at pH 9.0. A model of Est8 was constructed using a hypothetical YxiM precursor structure (2O14_A) from Bacillus subtilis as template. The structure showed an α/β-hydrolase fold and indicated the presence of a typical catalytic triad consisting of Ser-11, Asp-182, and His-185, which were investigated by site directed replacements coupled with kinetic characterization. Asp-182 and His-185 residues were more critical than the Ser-11 residue in the catalytic activity of Est8. A comparison of the amino acid sequence showed that Est8 could be grouped into the GDSL family and further classified as an SGNH hydrolase. Est8 is a new member of the SGNH hydrolase subfamily and may employ a different catalytic mechanism.
( Jun Pei Zhou ),( Yan Yan Dong ),( Jun Jun Li ),( Rui Zhang ),( Xianghua Tang ),( Yuelin Mu ),( Bo Xu ),( Qian Wu ),( Zun Xi Huang ) 한국미생물 · 생명공학회 2012 Journal of microbiology and biotechnology Vol.22 No.11
The α-galactosidase-coding gene agaAJB13 was cloned from Sphingomonas sp. JB13 showing 16S rDNA (1,343 bp) identities of ≤97.2% with other identified Sphingomonas strains. agaAJB13 (2,217 bp; 64.9% GC content) encodes a 738-residue polypeptide (AgaAJB13) with a calculated mass of 82.3 kDa. AgaAJB13 showed the highest identity of 61.4% with the putative glycosyl hydrolase family 36 α-galactosidase from Granulicella mallensis MP5ACTX8 (EFI56085). AgaAJB13 also showed <37% identities with reported protease-resistant or Sphingomonas α-galactosidases. A sequence analysis revealed different catalytic motifs between reported Sphingomonas α-galactosidases (KXD and RXXXD) and AgaAJB13 (KWD and SDXXDXXXR). Recombinant AgaAJB13 (rAgaAJB13) was expressed in Escherichia coli BL21 (DE3). The purified rAgaAJB13 was characterized using p-nitrophenyl-α-D-galactopyranoside as the substrate and showed an apparent optimum at pH 5.0 and 60oC and strong resistance to trypsin and proteinase K digestion. Compared with reported proteaseresistant α-galactosidases showing thermolability at 50oC or 60oC and specific activities of <71 U/mg with or without protease treatments, rAgaAJB13 exhibited a better thermal stability (half-life of >60 min at 60oC) and higher specific activities (225.0-256.5 U/mg). These sequence and enzymatic properties suggest AgaAJB13 is the first identified and characterized Sphingomonas α-galactosidase, and shows novel protease resistance with a potential value for basic research and industrial applications.
( Jun Pei Zhou ),( Qian Wu ),( Rui Zhang ),( Yu Ying Yang ),( Xiang Hua Tang ),( Jun Jun Li ),( Jun Mei Ding ),( Yan Yan Dong ),( Zun Xi Huang ) 한국미생물 · 생명공학회 2013 Journal of microbiology and biotechnology Vol.23 No.6
This paper reports the production and characterization of crude xylanase from the newly isolated Humicola sp. Ly01. The highest (41.8 U/ml) production of the crude xylanase was obtained under the optimized conditions (w/v): 0.5% wheat bran, 0.2% KH2PO4, and 0.5% peptone; initial pH 7.0; incubation time 72 h; 30℃; and 150 rpm. A considerable amount of the crude xylanase was induced using hulless barley bran or soybean meal as the carbon source, but a small amount of the enzyme was produced when supplementary urea was used as the nitrogen source to wheat bran. The crude xylanase showed apparent optimal cellulase-free xylanase activity at 60℃ and pH 6.0, more than 71.8% of the maximum xylanase activity in 3.0-30.0% (v/v) ethanol and more than 82.3% of the initial xylanase activity after incubation in 3.0-30.0% (v/v) ethanol at 30℃ for 2 h. The crude xylanase was moderately resistant to both acid and neutral protease digestion, and released 7.9 and 10.9 μmol/ml reducing sugar from xylan in the simulated gastric and intestinal fluids, respectively. The xylooligosaccharides were the main products of the hydrolysis of xylan by the crude xylanase. These properties suggested the potential of the crude enzyme for being applied in the animal feed industry, xylooligosaccharides production, and high-alcohol conditions such as ethanol production and brewing.
MULTIPLE SOLUTIONS FOR A p-LAPLACIAN SYSTEM WITH NONLINEAR BOUNDARY CONDITIONS
Zhou, Jun,Kim, Chan-Gyun Korean Mathematical Society 2014 대한수학회보 Vol.51 No.1
A nonlinear elliptic problem involving p-Laplacian and nonlinear boundary condition is considered in this paper. By using the method of Nehari manifold, it is proved that the system possesses two nontrivial nonnegative solutions if the parameter is small enough.
Development of action research in China: review and reflection
Jun Zhou,Katrina Yan Liua 서울대학교 교육연구소 2011 Asia Pacific Education Review Vol.12 No.2
Action research in China during late 1970s and 1980s was influenced by positivist scientific research, believing the purpose of doing action research is to explore the general law of education. It was carried out through educational experimentations emphasizing the control of the experimental conditions. Starting from 1990s, action research in China followed the positivist model of hypothesis verification on the one hand, while on the other hand, it introduced the notion of hermeneutics: finding and solving problems in real teaching situations and thereby understanding the real situation. Entering the new millennium, action research in China has further followed hermeneutics, accompanied by the emergence of "grassroots" action research.
Zhou, Can,Ren, Yu,Wang, Ke,Liu, Jie,He, Jian-Jun,Liu, Pei-Jun Asian Pacific Journal of Cancer Prevention 2013 Asian Pacific journal of cancer prevention Vol.14 No.12
Background and Objective: Rectal washout with saline solution may theoretically prevent anastomotic recurrence in patients with resectable rectal cancer, although exact clinical effects have not yet been determined. In order to derive a more precise estimation of the relationship, the present meta-analysis was performed. Method: Relevant studies were identified by a search of Medline, Embase and Google Website with no restrictions to September 1, 2013, and included in the systematic review and meta-analysis. Results: 5 trials (642 participants) were included to assess the association between rectal washout with saline solution and anastomotic recurrence. The rate of anastomotic recurrence (AR) was 6.23% (40/642), with the pooled OR derived from the five studies being 0.32 (95 % CI=0.15-0.70, P=0.004). The pooled OR derived from the TME and radical resection subgroups were 0.72 (95%CI=0.16-3.12, P=0.66) and 0.51 (95%CI=0.13-1.96, P=0.32), respectively. Conclusion: Results from this analysis show that intra-operative rectal washout with mere saline solution largely reduces the risk of anastomotic recurrences in patients with resectable rectal cancer.
Phylogenetic Diversity of Bacteria in an Earth-Cave in Guizhou Province, Southwest of China
Zhou, Jun-Pei,Gu, Ying-Qi,Zou, Chang-Song,Mo, Ming-He The Microbiological Society of Korea 2007 The journal of microbiology Vol.45 No.2
The objective of this study was to analyze the phylogenetic composition of bacterial community in the soil of an earth-cave (Niu Cave) using a culture-independent molecular approach. 16S rRNA genes were amplified directly from soil DNA with universally conserved and Bacteria-specific rRNA gene primers and cloned. The clone library was screened by restriction fragment length polymorphism (RFLP), and representative rRNA gene sequences were determined. A total of 115 bacterial sequence types were found in 190 analyzed clones. Phylogenetic sequence analyses revealed novel 16S rRNA gene sequence types and a high diversity of putative bacterial community. Members of these bacteria included Proteobacteria (42.6%), Acidobacteria (18.6%), Planctomycetes (9.0 %), Chloroflexi (Green nonsulfur bacteria, 7.5%), Bacteroidetes (2.1%), Gemmatimonadetes (2.7%), Nitrospirae (8.0%), Actinobacteria (High G+C Gram-positive bacteria, 6.4%) and candidate divisions (including the OP3, GN08, and SBR1093, 3.2%). Thirty-five clones were affiliated with bacteria that were related to nitrogen, sulfur, iron or manganese cycles. The comparison of the present data with the data obtained previously from caves based on 16S rRNA gene analysis revealed similarities in the bacterial community components, especially in the high abundance of Proteobacteria and Acidobacteria. Furthermore, this study provided the novel evidence for presence of Gemmatimonadetes, Nitrosomonadales, Oceanospirillales, and Rubrobacterales in a karstic hypogean environment.
Extraction and separation of D/L-lactic acid in simulated fermentation broth
Jun Zhou,노경호,Wentao Bi 한국화학공학회 2011 Korean Journal of Chemical Engineering Vol.28 No.7
The recovery of lactic acid from fermentation broth plays an important part in production of lactic acid. In this case, the extraction of lactic acid from simulated fermentation broth was processed by tri-n-octylamine dissolved in oleyl alcohol. The extraction efficiency was investigated with several variables, and the optimal condition of extraction of lactic acid (10 mg mL^−1) from aqueous solution was tri-n-octylamine/oleyl alcohol (30/70, v/v) and solvent phase/fermentation (1/2, v/v) stirred for 60 min under room temperature. The optimal back extraction of lactic acid was obtained by hot water (~90℃ ) with solvent phase/water (1/4, v/v). The back extracted racemic lactic acid was direct enantiomeric analyzed and separated by chiral ligand chromatography due to strict requirement of absolute configuration in pharmaceutical field and food science. The effect of various parameters on enantioselectivity was discussed and the (L)-phenylalaninamide·HCl (6.0 mmol L^−1) and CuSO^4·5H^2O (3.0 mmol L^−1) dissolved in methanol/water (5/95,v/v) at pH=6.0 was the suitable mobile phase for chiral ligand exchange separation of (D, L)-lactic acids. By the investigations,a convenient systemic method was established for extraction and separation of (D, L)-lactic acid.