RESUMO
Vanadium-based metallic salts, characterized by their intrinsic low electronic conductivity, are impeding their advancement as anode materials in the realm of lithium-ion battery technology. This study presents a novel embedded anode material KVO3/NC (KVO/NC) synthesized via a sol-gel method, with KVO3 (KVO) particles in situ growing on N-doped carbon, thereby ameliorating conductivity and electrochemical performance. The findings reveal that KVO/NC composite has three lithium-ion storage sites, ultra-high cycling stability (289 mA h/g@5000 cycles@10 C@100 %), and superior rate performance (249 mA h/g@15 C; 221 mA h/g@20 C). Coupled with LiFePO4 cathode, it achieves a competitive energy density (391 W h kg-1@0.1 C; 1-3.9 V). This work reveals the practical potential of KVO/NC as a new type of lithium-ion battery anode material with high energy density and long cycle life through a series of ex situ/in situ characterizations.
RESUMO
The antibiotic tetracycline (TC) is an emerging pollutant frequently detected in various environments. Biodegradation is a crucial approach for eliminating TC contamination. However, only a few efficient TC-degrading bacteria have been isolated, and the molecular mechanisms of TC degradation, as well as their application potential, remain poorly understood. This study isolated a novel TC-degrading bacterium, Providencia stuartii TX2, from the intestine of black soldier fly larvae. TX2 exhibited remarkable performance, degrading 72.17 % of 400 mg/L TC within 48 h. Genomic analysis of TX2 unveiled the presence of antibiotic resistance genes and TC degradation enzymes. Transcriptomic analysis highlighted the roles of proteins related to efflux pumps, enzymatic transformation, adversity resistance, and unknown functions. Three TC degradation pathways were proposed, with TC being transformed into 27 metabolites through epimerization, hydroxylation, oxygenation, ring opening, and de-grouping, reducing TC toxicity. Additionally, TX2 significantly enhanced TC biodegradation in four TC-contaminated environmental samples and reduced antibiotic resistance genes and mobile genetic elements in chicken manure. This research provides insights into the survival and biodegradation mechanisms of Providencia stuartii TX2 and evaluates its potential for environmental bioremediation.
Assuntos
Antibacterianos , Biodegradação Ambiental , Providencia , Tetraciclina , Providencia/genética , Providencia/metabolismo , Providencia/efeitos dos fármacos , Tetraciclina/metabolismo , Antibacterianos/metabolismo , Animais , Medição de Risco , Galinhas , Esterco/microbiologia , Larva/metabolismo , Larva/efeitos dos fármacosRESUMO
The widely used ET recombination requires an ssDNA product degraded by Rac phage protein E588 from dsDNA for strand invasion. However, proof of the ssDNA product is still elusive. The study provided three levels of proof sequentially. The probable ssDNAs degraded by E588 from the fluorescent plus-, minus-, or double-stranded dsDNA pET28a-xylanase exhibited a half fluorescence intensity of the corresponding dsDNAs, equivalent to the E588 degradation nucleotides half that of the total nucleotides degraded from the corresponding dsDNA. The ssDNA product degraded by E588 from the fluorescent minus-stranded dsDNA was confirmed by gradient gel-electrophoresis and two nuclease degradation reactions. Degraded by E588 from the dsDNA pET28a-xylanase that had a phosphorothioated plus-stranded 5'-terminus, the plus-stranded ssDNA product was separated via gel electrophoresis and recovered via a DNAclean kit. The recovered ssDNA product was proven to have intact 5'- and 3'-ends by DNA sequencing analysis. This study provides a solid foundation for the mechanism of ssDNA invasion.
RESUMO
Maize accumulates large amounts of starch in seeds which have been used as food for human and animals. Maize starch is an importantly industrial raw material for bioethanol production. One critical step in bioethanol production is degrading starch to oligosaccharides and glucose by α-amylase and glucoamylase. This step usually requires high temperature and additional equipment, leading to an increased production cost. Currently, there remains a lack of specially designed maize cultivars with optimized starch (amylose and amylopectin) compositions for bioethanol production. We discussed the features of starch granules suitable for efficient enzymatic digestion. Thus far, great advances have been made in molecular characterization of the key proteins involved in starch metabolism in maize seeds. The review explores how these proteins affect starch metabolism pathway, especially in controlling the composition, size and features of starch. We highlight the roles of key enzymes in controlling amylose/amylopectin ratio and granules architecture. Based on current technological process of bioethanol production using maize starch, we propose that several key enzymes can be modified in abundance or activities via genetic engineering to synthesize easily degraded starch granules in maize seeds. The review provides a clue for developing special maize cultivars as raw material in the bioethanol industry.
Assuntos
Amilose , Biocombustíveis , Etanol , Amido , Zea mays , Humanos , Amilopectina/metabolismo , Amilose/metabolismo , Engenharia Genética , Sementes/metabolismo , Amido/biossíntese , Amido/genética , Zea mays/genética , Zea mays/metabolismoRESUMO
In this study, the effect of corn oligopeptides (COPs) with liver protection activity on mice with hepatic fibrosis (HF) induced by carbon tetrachloride (CCl4 ) was studied. It was proved that COPs can ameliorate the liver injury and inflammation caused by CCl4 by histopathology and enzyme-linked immunosorbent assay in mice. The expression of Akt/NF-κB inflammatory pathway was determined by real-time polymerase chain reaction (RT-PCR) and western blotting (WB). The results showed that COPs inhibited the expression of key proteins in the inflammatory pathway. In conclusion, the results of this study suggested that COPs could improve CCl4 -induced HF by improving liver injury, reducing the expression of inflammatory factors, and inhibiting the expression of inflammatory signaling pathways. PRACTICAL APPLICATIONS: The corns around the world are mainly used as animal feed, and the liver protective activity of corn oligopeptides (COPs) is rarely applied to the market. The development of COPs liver protective food can prevent the occurrence of liver-related diseases such as hepatic fibrosis to a certain extent. Developing COPs liver protecting food can improve the utilization value of corn. It is hoped that this study can provide experimental support for the application of COPs in liver protection food.
Assuntos
NF-kappa B , Proteínas Proto-Oncogênicas c-akt , Animais , Cirrose Hepática/induzido quimicamente , Cirrose Hepática/tratamento farmacológico , Camundongos , NF-kappa B/genética , NF-kappa B/metabolismo , Oligopeptídeos/farmacologia , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais , Zea mays/metabolismoRESUMO
Because indel results in frame-shift mutations, seamless repair of double-stranded break (DSB)s plays a pivotal role in synthetic biology, molecular biology, and genome integrity. However, DSB repair is not well documented. T4 DNA ligase (T4lig) served to ligate intra-molecularly a zero bp break-apart DSB linear plasmid DNA pET22b(28a)-xylanase. An ATP T4lig ligation reaction joined one single-stranded break (SSB) into a phosphodiester-bond, whereas the opposite SSB into an abortive ligation intermediate blocking the DSB sequential repair. The intermediate proved to be fluorescent Cy5-AMP-SSB by a T4lig ligation reaction in the aid of Alexa Fluor 647 ATP having Cy5-AMP fluorescence. The fluorescent Cy5-AMP-SSB was de-adenylated into SSB by an ATP-free T4lig or Mg2+-free T4ligL159L reaction. The de-adenylated SSB was re-joined into another phosphodiester-bond by a sequential ATP T4lig re-ligation reaction. Thereby, DSB repair proceeds an abortive ligation, a reverse de-adenylation, and a sequential re-ligation reaction. The result has a potential usage in synthetic biology, molecular biology, and cancer-curing.
Assuntos
Trifosfato de Adenosina , Reparo do DNA , Monofosfato de Adenosina , PlasmídeosRESUMO
Developing for almost half a century, plasmid-construction has explored more than 37 methods. Some methods have evolved into new versions. From a global and evolutionary viewpoint, a review will make a clear understand and an easy practice for plasmid-construction. The 37 methods employ three principles as creating single-strand overhang, recombining homology arms, or serving amplified insert as mega-primer, and are classified into three groups as single strand overhang cloning, homologous recombination cloning, and mega-primer cloning. The methods evolve along a route for easy, efficient, or/and seamless cloning. Mechanism of plasmid-construction is primer annealing or/and primer invasion. Scar junction is a must-be faced scientific problem in plasmid-construction.
Assuntos
Recombinação Homóloga , Clonagem Molecular , Plasmídeos/genéticaRESUMO
BACKGROUND: Oil bodies (OBs) are a kind of natural and stable oil nucleate microcapsule in which the triglyceride matrix can be used as an appropriate carrier of hydrophobic molecules. Astaxanthin has high antioxidant properties but is extremely sensitive to oxidation, causing the loss of its bioactive properties. RESULTS: The purpose of this study was to clarify the effects of environmental factors (light, oxygen, temperature, and pH) on the physical and oxidative stability of astaxanthin microcapsules prepared with peanut oil bodies (POBs). After 14 days of storage, the retention rate of astaxanthin in peanut oil microcapsules (POMs) was significantly increased. The astaxanthin retention rate of POMs stored under light conditions was higher than under dark conditions. Similarly, the retention rate of astaxanthin in POMs was significantly increased during vacuum storage. The astaxanthin retention rate was also the highest when POMs were stored at 4 °C, whereas it was the lowest at pH 3.0. CONCLUSION: The experiment demonstrated that microcapsulation could improve the astaxanthin retention rate and storage stability, and recombinant OBs were potential ideal wall materials for astaxanthin embedding. © 2022 Society of Chemical Industry.
Assuntos
Lipossomos , Xantofilas , Cápsulas/química , Estresse Oxidativo , Xantofilas/químicaRESUMO
Although many methods have been reported, plasmid construction compromises transformant efficiency (number of transformants per ng of DNAs) with plasmid accuracy (rate of scarless plasmids). An efficient method is two-step PCR serving DNA amplification. An accurate method is ExnaseII cloning serving homology recombination (HR). We combine DNA amplification and HR to develop an intra-molecular HR by amplifying plasmid DNAs to contain homology 5'- and 3'-terminus and recombining the plasmid DNAs in vitro. An example was to construct plasmid pET20b-AdD. The generality was checked by constructing plasmid pET21a-AdD and pET22b-AdD in parallel. The DNAs having 30-bp homology arms were optimal for intra-molecular HR, and transformation of which created 14.2 transformants/ng and 90% scarless plasmids, more than the two-step PCR and the ExnaseII cloning. Transformant efficiency correlated with the component of nicked circular plasmid DNAs of HR products, indicating nick modification in vivo leads to scar plasmids.
Assuntos
Engenharia Genética/métodos , Recombinação Homóloga , Plasmídeos/genética , Clonagem Molecular/métodos , Escherichia coli , Transformação BacterianaRESUMO
Used as model for sandwich fusion, a mesophilic Aspergillus niger GH11 xylanase (Xyn) was fused into C2-Xyn-C2 with a thermophilic Thermotaga maritima GH10 xylanase carbohydrate-binding module CBM9_2 (C2). Linearized plasmids C2-pET20b-C2-Xyn were amplified from template pET20b-Xyn-C2 with a 4.3â¯kb C2-pET20b megaprimer, ligated into circular plasmids in blunt-end ligation, and transformed into E. coli BL21 (DE3) cells. The C2-Xyn-C2 had optimum activity at 45⯰C and pHâ¯4.2, a 2.85â¯h thermal inactivation half-life at 80⯰C and a 8.69â¯h at 50⯰C, with the 8.69â¯h value 24.8-, 7.5-, and 7.1-fold longer than the Xyn and single terminal fusion enzymes Xyn-C2, and C2-Xyn. Thermodynamics showed that the enzyme had a 1.8⯰C higher melting temperature, lower values ΔS, ΔΔG, and a denser structure than the Xyn. Kinetics showed that the C2-Xyn-C2 catalytic efficiency was 1.2-~6-fold and 2.7-~7.9-fold higher on beechwood and oat-spelt xylan than those of the enzymes Xyn, Xyn-C2, and C2-Xyn. The sandwich fusion evolved the xylanase with "armor-hands" to enhance simultaneously thermostability and activity in quality.
Assuntos
Aspergillus niger/enzimologia , Bactérias/enzimologia , Endo-1,4-beta-Xilanases/genética , Proteínas Recombinantes de Fusão/genética , Sequência de Aminoácidos , Endo-1,4-beta-Xilanases/química , Estabilidade Enzimática , Escherichia coli/genética , Temperatura Alta , Cinética , Proteínas Recombinantes de Fusão/química , Especificidade por SubstratoRESUMO
There exist significant differences between the 2 main types of xylanases, family F10 and G11. A clear understanding of the expression pattern of microbial F10 and G11 under different culture conditions would facilitate better production and industrial application of xylanase. In this study, the fungal xylanase producer Aspergillus niger A09 was systematically investigated in terms of induced expression of xylanase F10 and G11. Results showed that carbon and nitrogen sources could influence xylanase F10 and G11 transcript abundance, with G11 more susceptible to changes in culture media composition. The most favorable carbon and nitrogen sources for high G11 and low F10 production by A. niger A09 were xylan (2%) and (NH4)2C2O4 (0.3%), respectively. Following cultivation at 33 °C for 60 h, the highest xylanase activity (1132 IU per gram of wet mycelia) was observed. On the basis of differential gene expression of F10 and G11, as well as their different properties, we deduced that the F10 protein initially targeted xylan and hydrolyzed it into fragments including xylose, after which xylose acted as the inducer of F10 and G11 gene expression. These speculations also accounted for our failure to identify conditions favoring the high production of F10 but a low production of G11.
Assuntos
Aspergillus niger/enzimologia , Endo-1,4-beta-Xilanases/biossíntese , Carbono/metabolismo , Hidrólise , Nitrogênio/metabolismo , Reação em Cadeia da Polimerase em Tempo RealRESUMO
OBJECTIVE: The 9_2 carbohydrate-binding module (C2) locates natively at the C-terminus of the GH10 thermophilic xylanase from Thermotoga marimita. When fused to the C-terminus, C2 improved thermostability of a GH11 xylanase (Xyn) from Aspergillus niger. However, a question is whether the C-terminal C2 would have a thermostabilizing effect when fused to the N-terminus of a catalytic module. RESULTS: A chimeric enzyme, C2-Xyn, was created by step-extension PCR, cloned in pET21a(+), and expressed in E. coli BL21(DE3). The C2-Xyn exhibited a 2 °C higher optimal temperature, a 2.8-fold longer thermostability, and a 4.5-fold higher catalytic efficiency on beechwood xylan than the Xyn. The C2-Xyn exhibited a similar affinity for binding to beechwood xylan and a higher affinity for oat-spelt xylan than Xyn. CONCLUSION: C2 is a thermostabilizing carbohydrate-binding module and provides a model of fusion at an enzymatic terminus inconsistent with the modular natural terminal location.
Assuntos
Aspergillus niger/enzimologia , Endo-1,4-beta-Xilanases/química , Endo-1,4-beta-Xilanases/genética , Thermotoga maritima/enzimologia , Aspergillus niger/química , Aspergillus niger/genética , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Domínio Catalítico , Endo-1,4-beta-Xilanases/metabolismo , Estabilidade Enzimática , Proteínas Fúngicas/química , Proteínas Fúngicas/genética , Engenharia de Proteínas , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Especificidade por Substrato , Thermotoga maritima/química , Thermotoga maritima/genéticaRESUMO
The Aspergillus niger xylanase (Xyn) was used as a model to investigate impacts of un-structured residues on GH11 family enzyme, because the ß-jelly roll structure has five residues (Ser1Ala2Gly3Ile4Asn5) at N-terminus and two residues (Ser183Ser184) at C-terminus that do not form to helix or strand. The N- or/and C-terminal residues were respectively deleted to construct three mutants. The optimal temperatures of XynΔN, XynΔC, and XynΔNC were 46, 50, and 46°C, and the thermostabilities were 15.7, 73.9, 15.5 min at 50°C, respectively, compared to 48°C and 33.9 min for the Xyn. After kinetic analysis, the substrate-binding affinities for birch-wood xylan decreased in the order XynΔC>Xyn>XynΔNC>XynΔN, while the K(cat) values increased in the order XynΔCAssuntos
Aspergillus niger/enzimologia
, Endo-1,4-beta-Xilanases/química
, Sequência de Aminoácidos
, Domínio Catalítico
, Endo-1,4-beta-Xilanases/genética
, Estabilidade Enzimática
, Concentração de Íons de Hidrogênio
, Cinética
, Modelos Moleculares
, Dados de Sequência Molecular
, Ligação Proteica
, Estrutura Secundária de Proteína
, Deleção de Sequência
, Especificidade por Substrato
RESUMO
BACKGROUND: Domain fusion is limited at enzyme one terminus. The issue was explored by swapping a mesophilic Aspergillus niger GH11 xylanase (Xyn) with a hyper-thermophilic Thermotoga maritima glucanase (Glu) to construct two chimeras, Xyn-Glu and Glu-Xyn, with an intention to create thermostable xylanase containing glucanase activity. RESULTS: When expressed in E. coli BL21(DE3), the two chimeras exhibited bi-functional activities of xylanase and glucanase. The Xyn-Glu Xyn moiety had optimal reaction temperature (Topt) at 50 °C and thermal in-activation half-life (t1/2) at 50 °C for 47.6 min, compared to 47 °C and 17.6 min for the Xyn. The Glu-Xyn Xyn moiety had equivalent Topt to and shorter t1/2 (5.2 min) than the Xyn. Both chimera Glu moieties were more thermostable than the Glu, and the three enzyme Topt values were higher than 96 °C. The Glu-Xyn Glu moiety optimal pH was 5.8, compared to 3.8 for the Xyn-Glu Glu moiety and the Glu. Both chimera two moieties cooperated with each other in degrading substrates. CONCLUSIONS: Domain-swapping created different effects on each moiety properties. Fusing the Glu domain at C-terminus increased the xylanase thermostability, but fusing the Glu domain at N-terminus decreased the xylanase thermostability. Fusing the Xyn domain at either terminus increased the glucanase thermostability, and fusing the Xyn domain at C-terminus shifted the glucanase pH property 2 units higher towards alkaline environments. Fusing a domain at C-terminus contributes more to enzyme catalytic activity; whereas, fusing a bigger domain at N-terminus disturbs enzyme substrate binding affinity.
Assuntos
Dextranase/metabolismo , Endo-1,4-beta-Xilanases/metabolismo , Sequência de Aminoácidos , Aspergillus niger/enzimologia , Dextranase/química , Dextranase/genética , Endo-1,4-beta-Xilanases/química , Endo-1,4-beta-Xilanases/genética , Escherichia coli/metabolismo , Meia-Vida , Concentração de Íons de Hidrogênio , Cinética , Dados de Sequência Molecular , Estrutura Terciária de Proteína , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Temperatura , Thermotoga maritima/enzimologiaRESUMO
Because the genetic codon is known for degeneracy, its effect on enzyme thermal property is seldom investigated. A dataset was constructed for GH10 xylanase coding sequences and optimal temperatures for activity (T(opt)). Codon contents and relative synonymous codon usages were calculated and respectively correlated with the enzyme T(opt) values, which were used to describe the xylanase thermophilic tendencies without dividing them into two thermophilic and mesophilic groups. After analyses of codon content and relative synonymous codon usages were checked by the Bonferroni correction, we found five codons, with three (AUA, AGA, and AGG) correlating positively and two (CGU and AGC) correlating negatively with the T(opt) value. The three positive codons are purine-rich codons, and the two negative codons have A-ends. The two negative codons are pyridine-rich codons, and one has a C-end. Comparable with the codon C- and A-ending features, C- and A-content within mRNA correlated negatively and positively with the T(opt) value, respectively. Thereby, codons have effects on enzyme thermal property. When the issue is analyzed at the residual level, the effect of codon message is lost. The codons relating to enzyme thermal property are selected by thermophilic force at nucleotide level.
Assuntos
Códon , Xilosidases/metabolismo , RNA Mensageiro/genéticaRESUMO
Protein structure is composed of regular secondary structural elements (α-helix and ß-strand) and non-regular region. Unlike the helix and strand, the non-regular region consists of an amino acid defined as a disordered residue (DR). When compared with the effect of the helix and strand, the effect of the DR on enzyme structure and function is elusive. An Aspergillus niger GH10 xylanase (Xyn) was selected as a model molecule of (ß/α)(8) because the general structure consists of ~10% enzymes. The Xyn has five N-terminal DRs and one C-terminal DR, respectively, which were deleted to construct three mutants, XynΔN, XynΔC, and XynΔNC. Each mutant was ~2-, 3-, or 4-fold more thermostable and 7-, 4-, or 4-fold more active than the Xyn. The N-terminal deletion decreased the xylanase temperature optimum for activity (T(opt)) 6 °C, but the C-terminal deletion increased its T(opt) 6 °C. The N- and C-terminal deletions had opposing effects on the enzyme T(opt) but had additive effects on its thermostability. The five N-terminal DR deletions had more effect on the enzyme kinetics but less effect on its thermo property than the one C-terminal DR deletion. CD data showed that the terminal DR deletions increased regular secondary structural contents, and hence, led to slow decreased Gibbs free energy changes (ΔG(0)) in the thermal denaturation process, which ultimately enhanced enzyme thermostabilities.
Assuntos
Aspergillus niger/enzimologia , Endo-1,4-beta-Xilanases/química , Proteínas Fúngicas/química , Modelos Moleculares , Sequência de Aminoácidos , Aspergillus niger/genética , Endo-1,4-beta-Xilanases/genética , Proteínas Fúngicas/genética , Estrutura Secundária de Proteína , Estrutura Terciária de Proteína , Deleção de SequênciaRESUMO
Besides the catalytic domain, some xylanases contained a non-catalytic domain which is named as carbohydrate binding module (CBM). CBM can be used to improve their binding-ability to insoluble substrates. We illustrated the importance of CBM by reviewing the source of CBMs, type of families, features of binding to insoluble substrates, specific amino acids involved in substrate-binding, linker peptides connecting the catalytic domain, and the effect of CBMs on xylanase thermostability. CBM is important for xylanase to break down complicate carbohydrates. Perspectives on engineering xylanase activity according to the characteristics of CBMs were given.
Assuntos
Metabolismo dos Carboidratos , Endo-1,4-beta-Xilanases/metabolismo , Sítios de Ligação , Catálise , Complexos Multienzimáticos/química , Especificidade por SubstratoRESUMO
Dialdehyde starch (DAS) was used as a novel coupling agent to prepare chitosan carrier to immobilize the xylanase from Aspergillus niger A-25. Compared with glutaraldehyde-cross-linked chitosan (CS-GA) and pure chitosan beads, the DAS-cross-linked chitosan (CS-DAS) beads exhibited the highest xylanase activity recovery. The DAS adding amount and cross-linking time in CS-DAS preparation process were optimized with respect to activity recovery to the values of 1.0 g (6.7% w/v concentration) and 16 h, respectively. The optimum temperature of both the CS-DAS- and CS-GA-immobilized xylanase was observed to be 5 degrees C higher than that of free enzyme (50 degrees C). The CS-DAS-immobilized xylanase had the highest thermal and storage stability as compared to the CS-GA-immobilized and free xylanase. The apparent K (m) and V (max) values of the CS-DAS-immobilized xylanase were estimated to be 1.29 mg/ml and 300.7 mumol/min/mg protein, respectively. The CS-DAS-immobilized xylanase could produce from birchwood xylan high-quality xylo-oligosaccharides, mainly composed of xylotriose, as free xylanase did. The proposed CS-DAS carrier was more advantageous over the CS-GA or pure chitosan carrier for xylanase immobilization application.
Assuntos
Aspergillus niger/enzimologia , Quitosana/química , Reagentes de Ligações Cruzadas/química , Endo-1,4-beta-Xilanases/química , Enzimas Imobilizadas/química , Amido/análogos & derivados , Betula/química , Biocatálise , Materiais Biocompatíveis/química , Endo-1,4-beta-Xilanases/metabolismo , Estabilidade Enzimática , Enzimas Imobilizadas/metabolismo , Cinética , Amido/química , Fatores de Tempo , Xilanos/metabolismoRESUMO
A bioinformatics method was used to search the responsible dipeptides for optimum pH in the G/11 xylanase, for dipeptides can provide position information of the related residues for rational protein design. The responsible dipeptides were found as negative YS and positive SY, GR, MR, and KR. The minimum and maximum optimum pH was calculated as 2.33 and 14.29, respectively. Compared with the known crystal structures of the G/11 xylanase, YS was found mostly in the turn area of beta-strands of S/T surface; and SY was found in the inner part of beta-strands of the S/T area near to the active site of proton donor; and the GR, MR, and KR in the coil region connecting "finger" to the alpha-helix. The result clearly explained the success of shifting of pH 0.5 U to alkaline by the introduction of arginines into S/T area of a xylanase. The result would be useful for xylanase engineering, and the adaptation mechanism to high alkaline was also discussed.
Assuntos
Biologia Computacional , Dipeptídeos/química , Dipeptídeos/metabolismo , Endo-1,4-beta-Xilanases/química , Endo-1,4-beta-Xilanases/metabolismo , Aspergillus/enzimologia , Bacillus/enzimologia , Sítios de Ligação , Simulação por Computador , Concentração de Íons de Hidrogênio , Modelos Moleculares , Estrutura Terciária de ProteínaRESUMO
A bioinformatics method was used to analyze F/10 and G/11 xylanase basing on principle component analysis, and a model was made to classify between these two folds with an ideal result. The principle components were predicated to be secondary structures, the components were analyzed with the architecture of each family, and found comparable with (beta/alpha)(8)-barrel of F/10 xylanase and right-hand structure of G/11 xylanase. Compared with sequence similarities, this method gave discriminating features a clear meaning. The largest component did not appear in the model, which revealed no difference between these two families.