白色链霉菌乙酰木聚糖酯酶基因的克隆及生物信息学分析

乙酰木聚糖酯酶可以水解乙酰化木聚糖中的O-乙酰取代基团,消除该基团对木聚糖酶水解的空间阻碍作用,增强木聚糖酶对木聚糖的亲和力和降解能力。以白色链霉菌基因组为模板,利用简并PCR和TAIL-PCR扩增获得长约741 bp阅读框片段,编码247个氨基酸。生物信息学分析表明,该多肽片段具有AXE1家族蛋白保守区域;与已知的乙酰木聚糖酯酶蛋白C端区相比,相似性较高,二级和三级结构空间排布特点极为相似;初步判定该多肽片段为白色链霉菌乙酰木聚糖酯酶的C端区域。     

Molecular cloning, and characterization of a modular acetyl xylan esterase from the edible straw mushroom Volvariella volvacea

A new Volvariella volvacea gene encoding an acetyl xylan esterase (designated as Vvaxe1) was cloned and expressed in Pichia pastoris. The cDNA contained an ORF of 1047 bp encoding 349 amino acids with a calculated mass of 39 990 Da. VvAXE1 is a modular enzyme consisting of an N-terminal signal peptide, a catalytic domain, and a cellulose-binding domain. The amino acid sequence of the enzyme exhibited a high degree of similarity to cinnamoyl esterase B from Penicillium funiculosum, and acetyl xylan esterases from Aspergillus oryzae, Penicillium purpurogenum, and Aspergillus ficuum. Recombinant acetyl xylan esterase released acetate from several acetylated substrates including beta-d-xylose tetraacetate and acetylated xylan. No activity was detectable on p-nitrophenyl acetate. Enzyme-catalyzed hydrolysis of 4-methylumbelliferyl acetate was maximal at pH 8.0 and 60 degrees C, and reciprocal plots revealed an apparent K(m) value of 307.7 microM and a V(max) value of 24 733 IU micromol(-1) protein. ReAXE1 also exhibited a capacity to bind to Avicel and H(3)PO(4) acid-swollen cellulose.     

The crystal structure of the cephalosporin deacetylating enzyme acetyl xylan esterase bound to paraoxon explains the low sensitivity of this serine hydrolase to organophosphate inactivation

Organophosphorus insecticides and nerve agents irreversibly inhibit serine hydrolase superfamily enzymes. One enzyme of this superfamily, the industrially important (for β-lactam antibiotic synthesis) AXE/CAH (acetyl xylan esterase/cephalosporin acetyl hydrolase) from the biotechnologically valuable organism Bacillus pumilus, exhibits low sensitivity to the organophosphate paraoxon (diethyl-p-nitrophenyl phosphate, also called paraoxon-ethyl), reflected in a high K(i) for it (~5 mM) and in a slow formation (t(?)~1 min) of the covalent adduct of the enzyme and for DEP (E-DEP, enzyme-diethyl phosphate, i.e. enzyme-paraoxon). The crystal structure of the E-DEP complex determined at 2.7 ? resolution (1 ?=0.1 nm) reveals strain in the active Ser1?1-bound organophosphate as a likely cause for the limited paraoxon sensitivity. The strain results from active-site-size limitation imposed by bulky conserved aromatic residues that may exclude as substrates esters having acyl groups larger than acetate. Interestingly, in the doughnut-like homohexamer of the enzyme, the six active sites are confined within a central chamber formed between two 60°-staggered trimers. The exclusive access to this chamber through a hole around the three-fold axis possibly limits the size of the xylan natural substrates. The enzyme provides a rigid scaffold for catalysis, as reflected in the lack of movement associated with paraoxon adduct formation, as revealed by comparing this adduct structure with that also determined in the present study at 1.9 ? resolution for the paraoxon-free enzyme.     

Deacetylation of xylans by acetyl esterases of Trichoderma reesei

Summary Two previously purified esterases of Trichoderma reesei were used to study the deacetylation of polymeric, oligomeric and dimeric acetylated xylan fragments. For the first time nearly complete enzymatic deacetylation of polymeric xylan with purified acetyl xylan esterase was demonstrated, resulting in precipitation of the remaining polymer structure. The esterases had very different substrate specifities, one having a preference for high molecular weight substrates and the other showing high activity only towards acetyl xylobiose. The latter enzyme was also regioselective, cleaving off the acetyl substituent only from the C-3 position of the xylopyranose ring. The highest xylose yield from acetylated xylan was obtained by the synergistic action of xylanase, \-xylosidase and acetyl xylan esterase. Offprint requests to: M. Sundberg     

植物根际芽胞杆菌菌种资源采集及其具有过水解催化活性水解酶基因的克隆

[背景]芽胞杆菌源枯草杆菌蛋白酶(subtilisin carlsberg)、乙酰基木聚糖酯酶(acetyl xylan esterase)和头孢菌素乙酰水解酶(cephalosporin acetyl hydrolase)具有较高的过水解催化活性,有商业开发价值。[目的]挖掘芽胞杆菌菌株中具有过水解酶催化活性的水解酶蛋白基因,为后续制备过水解酶及酶法合成过氧乙酸奠定基础。[方法]利用定向筛选培养基,从植物根际及纳豆产品中筛选产蛋白酶芽胞杆菌候选菌株,并利用RFLP及16S rRNA基因对其进行鉴定。从蛋白酶高产芽胞杆菌菌株中克隆枯草杆菌蛋白酶、乙酰木聚糖醋酶和头孢菌素乙酰水解酶的全长基因。[结果]从植物根际土壤及纳豆产品中共分离到85个候选菌株,RFLP及16S rRNA基因鉴定结果表明候选菌株均为芽胞杆菌,分别属于Bacillus subtilis、Bacillus cereus、Bacillus pumilus和Bacillus megaterium四个类群。从B.subtilis NSYT-3克隆的枯草杆菌蛋白酶基因编码的多肽链全长381个氨基酸,从B.pumilus OSLJ-3克隆得到的乙酰基木聚糖酯酶基因编码的多肽链全长320个氨基酸,从B.subtilis NSYT-3克隆的头孢菌素乙酰水解酶基因编码的多肽链全长318个氨基酸,3D结构模拟表明这3个酶蛋白均具有α/β水解酶折叠家族蛋白结构特点。[结论]芽胞杆菌源具过水解催化活性水解酶基因的克隆,为后续开发酶法合成过氧乙酸工艺奠定了基础。     

Hyperthermostable acetyl xylan esterase

An esterase which is encoded within a Thermotoga maritima chromosomal gene cluster for xylan degradation and utilization was characterized after heterologous expression of the corresponding gene in Escherichia coli and purification of the enzyme. The enzyme, designated AxeA, shares amino acid sequence similarity and its broad substrate specificity with the acetyl xylan esterase from Bacillus pumilus, the cephalosporin C deacetylase from Bacillus subtilis, and other (putative) esterases, allowing its classification as a member of carbohydrate esterase family 7. The recombinant enzyme displayed activity with p-nitrophenyl-acetate as well as with various acetylated sugar substrates such as glucose penta-acetate, acetylated oat spelts xylan and DMSO (dimethyl sulfoxide)-extracted beechwood xylan, and with cephalosporin C. Thermotoga maritima AxeA represents the most thermostable acetyl xylan esterase known to date. In a 10 min assay at its optimum pH of 6.5 the enzyme's activity peaked at 90°C. The inactivation half-life of AxeA at a protein concentration of 0.3 µg µl⁻¹ in the absence of substrate was about 13 h at 98°C and about 67 h at 90°C. Differential scanning calorimetry analysis of the thermal stability of AxeA corroborated its extreme heat resistance. A multi-phasic unfolding behaviour was found, with two apparent exothermic peaks at approximately 100–104°C and 107.5°C. In accordance with the crystal structure, gel filtration analysis at ambient temperature revealed that the enzyme has as a homohexameric oligomerization state, but a dimeric form was also found.     

Characterization of an acetyl xylan esterase from the anaerobic fungus Orpinomyces sp. strain PC-2.

A 1,067-bp cDNA, designated axeA, coding for an acetyl xylan esterase (AxeA) was cloned from the anaerobic rumen fungus Orpinomyces sp. strain PC-2. The gene had an open reading frame of 939 bp encoding a polypeptide of 313 amino acid residues with a calculated mass of 34,845 Da. An active esterase using the original start codon of the cDNA was synthesized in Escherichia coli. Two active forms of the esterase were purified from recombinant E. coli cultures. The size difference of 8 amino acids was a result of cleavages at two different sites within the signal peptide. The enzyme released acetate from several acetylated substrates, including acetylated xylan. The activity toward acetylated xylan was tripled in the presence of recombinant xylanase A from the same fungus. Using p-nitrophenyl acetate as a substrate, the enzyme had a K(m) of 0.9 mM and a V(max) of 785 micromol min(-1) mg(-1). It had temperature and pH optima of 30 degrees C and 9.0, respectively. AxeA had 56% amino acid identity with BnaA, an acetyl xylan esterase of Neocallimastix patriciarum, but the Orpinomyces AxeA was devoid of a noncatalytic repeated peptide domain (NCRPD) found at the carboxy terminus of the Neocallimastix BnaA. The NCRPD found in many glycosyl hydrolases and esterases of anaerobic fungi has been postulated to function as a docking domain for cellulase-hemicellulase complexes, similar to the dockerin of the cellulosome of Clostridium thermocellum. The difference in domain structures indicated that the two highly similar esterases of Orpinomyces and Neocallimastix may be differently located, the former being a free enzyme and the latter being a component of a cellulase-hemicellulase complex. Sequence data indicate that AxeA and BnaA might represent a new family of hydrolases.     

A combined approach for improving alkaline acetyl xylan esterase production in Pichia pastoris, and effects of glycosylation on enzyme secretion, activity and stability

High level expression of axe1, a gene previously cloned from Volvariella volvacea that encodes an acetyl xylan esterase with two potential N-linked glycosylation sites, has been achieved in Pichia pastoris using a codon-optimized axe1 synthesized by the primer extension PCR procedure. The GC content of the codon-optimized axe1 was 48.62% compared with 55.49% in the native gene. Using the codon-optimized construct, AXE1 expression in P. pastoris was increased from an undetectable level to 136.45U/ml six days after induction of yeast cultures grown in BMMY medium. A further increase (to 463U/ml) was achieved when conditions for yeast culture were optimized as follows: 2.8% methanol, 0.63% casamino acids, and pH 8.0. This latter value represented a 3.4-fold and 246-fold increase in the enzyme levels recorded in non-optimized P. pastoris cultures and in rice straw-grown cultures of V. volvacea, respectively. N-linked glycosylation played an essential role in AXE1 secretion but had only a slight effect on the catalytic activity and stability of the recombinant enzyme.     

A novel cephalosporin deacetylating acetyl xylan esterase from Bacillus subtilis with high activity toward cephalosporin C and 7-aminocephalosporanic acid

A cephalosporin deacetylating acetyl xylan esterase was cloned from the genomic DNA of Bacillus subtilis CICC 20034 and functionally expressed in Escherichia coli. Its gene contained an open reading frame of 957 bp encoding 318 amino acids with a calculated mass of 35,607 Da, and it displayed significant identity to acetyl xylan esterases from Bacillus sp. 916, B. subtilis 168, and Bacillus pumilus Cect5072. The enzyme was a native homohexamer but a trimer under the condition of 1 % sodium dodecyl sulfate (SDS); both forms were active and could transit to each other by incubating in or removing SDS. The enzyme belongs to carbohydrate esterase family 7 and had a double specificity on both the acetylated oligosaccharide and cephalosporin C (CPC) and 7-aminocephalosporanic acid (7-ACA). The activity of this purified enzyme toward CPC and 7-ACA was highest among all the acetyl xylan esterase from CE family 7, which were 484 and 888 U/mg, respectively, and endowed itself with great industrial interest on semi-synthetic β-lactam antibiotics. The optimum pH of the purified enzyme was 8.0, and the optimum temperature was 50 °C, and the enzyme had high thermal stability, broad range of pH tolerance, and extremely organic solvent tolerance.     

Carbohydrate esterase family 4 enzymes: substrate specificity

The substrate specificity of selected enzymes classified under Carbohydrate Esterase family 4 (CE4) has been examined. Chitin deacetylase from Mucor rouxii and both a native and a truncated form of acetyl xylan esterase from Streptomyces lividans were found to be active on both xylan and several soluble chitinous substrates. Furthermore, the activities of all enzymes examined were significantly increased in the presence of Co(2+) when chitinous substrates were employed. However, the presence of this metal ion did not result in enhancing the activities of the enzymes when xylan was used as substrate. An acetyl xylan esterase from Bacillus pumilus, classified under Carbohydrate Esterase family 7, was found to be inactive towards all chitinous substrates tested. Finally, all enzymes examined were inactive towards cell wall peptidoglycan.     

Penicillium purpurogenum produces a family 1 acetyl xylan esterase containing a carbohydrate-binding module: characterization of the protein and its gene

At least three acetyl xylan esterases (AXE I, II and III) are secreted by Penicillium purpurogenum. This publication describes more detailed work on AXE I and its gene. AXE I binds cellulose but not xylan; it is glycosylated and inactivated by phenylmethylsulphonyl fluoride, showing that it is a serine esterase. The axe1 gene presents an open reading frame of 1278 bp, including two introns of 68 and 61 bp; it codes for a signal peptide of 31 residues and a mature protein of 351 amino acids (molecular weight 36,693). AXE I has a modular structure: a catalytic module at the amino terminus belonging to family 1 of the carbohydrate esterases, a linker rich in serines and threonines, and a family 1 carboxy terminal carbohydrate binding module (CBM). The CBM is similar to that of AXE from Trichoderma reesei, (with a family 5 catalytic module) indicating that the genes for catalytic modules and CBMs have evolved separately, and that they have been linked by gene fusion. The promoter sequence of axe1 contains several putative sequences for binding of gene expression regulators also found in other family 1 esterase gene promoters. It is proposed that AXE I and II act in succession in xylan degradation; first, xylan is attacked by AXE I and other xylanases possessing CBMs (which facilitate binding to lignocellulose), followed by other enzymes acting mainly on soluble substrates.     

Distinct roles of carbohydrate esterase family CE16 acetyl esterases and polymer-acting acetyl xylan esterases in xylan deacetylation

Mass spectrometric analysis was used to compare the roles of two acetyl esterases (AE, carbohydrate esterase family CE16) and three acetyl xylan esterases (AXE, families CE1 and CE5) in deacetylation of natural substrates, neutral (linear) and 4-O-methyl glucuronic acid (MeGlcA) substituted xylooligosaccharides (XOS). AEs were similarly restricted in their action and apparently removed in most cases only one acetyl group from the non-reducing end of XOS, acting as exo-deacetylases. In contrast, AXEs completely deacetylated longer neutral XOS but had difficulties with the shorter ones. Complete deacetylation of neutral XOS was obtained after the combined action of AEs and AXEs. MeGlcA substituents partially restricted the action of both types of esterases and the remaining acidic XOS were mainly substituted with one MeGlcA and one acetyl group, supposedly on the same xylopyranosyl residue. These resisting structures were degraded to great extent only after inclusion of α-glucuronidase, which acted with the esterases in a synergistic manner. When used together with xylan backbone degrading endoxylanase and β-xylosidase, both AE and AXE enhanced the hydrolysis of complex XOS equally.     

Isolation, analysis, and expression of two genes from Thermoanaerobacterium sp. strain JW/SL YS485: a beta-xylosidase and a novel acetyl xylan esterase with cephalosporin C deacetylase activity.

The genes encoding acetyl xylan esterase 1 (axe1) and a beta-xylosidase (xylB) have been cloned and sequenced from Thermoanaerobacterium sp. strain JW/SL YS485. axe1 is located 22 nucleotides 3' of the xylB sequence. The identity of axe1 was confirmed by comparison of the deduced amino acid sequence to peptide sequence analysis data from purified acetyl xylan esterase 1. The xylB gene was identified by expression cloning and by sequence homology to known beta-xylosidases. Plasmids which independently expressed either acetyl xylan esterase 1 (pAct1BK) or beta-xylosidase (pXylo-1.1) were constructed in Escherichia coli. Plasmid pXylAct-1 contained both genes joined at a unique EcoRI site and expressed both activities. Substrate specificity, pH, and temperature optima were determined for partially purified recombinant acetyl xylan esterase 1 and for crude recombinant beta-xylosidase. Similarity searches showed that the axe1 and xylB genes were homologs of the ORF-1 and xynB genes, respectively, isolated from Thermoanaerobacterium saccharolyticum. Although the deduced sequence of the axe1 product had no significant amino acid sequence similarity to any reported acetyl xylan esterase sequence, it did have strong similarity to cephalosporin C deacetylase from Bacillus subtilis. Recombinant acetyl xylan esterase 1 was found to have thermostable deacetylase activity towards a number of acetylated substrates, including cephalosporin C and 7-aminocephalosporanic acid.     

Regioselective deacetylation of cellulose acetates by acetyl xylan esterases of different CE-families

Cellulose acetate (CA) was found to be a substrate of several acetyl xylan esterases (AXE). Eight AXE from different carbohydrate esterase (CE) families were tested on their activity against CA with a degree of substitution of 0.7 and 1.4. The classification of the AXEs into CE families according to their structure by hydrophobic cluster analysis followed clearly their activity against CA. Within the same CE family similar, and between the CE families different deacetylation behaviours could be observed. Furthermore, each esterase family showed a distinct regioselective mode of action. The CE 1 family enzymes regioselectively cleaved the substituents in C2- and C3-position, while CE 5 family enzymes only cleaved the acetyl groups in C2-position. CE 4 family enzymes seemed to interact only with the substituents in C3-position. Evidence was found that the deacetylation reaction of the CE 1 family enzymes proceeded faster in C2- than in C3-position of CA. The enzymes were able to cleave acetyl groups from fully substituted anhydroglucose units.     

Expression of fungal acetyl xylan esterase in Arabidopsis thaliana improves saccharification of stem lignocellulose

Cell wall hemicelluloses and pectins are O‐acetylated at specific positions, but the significance of these substitutions is poorly understood. Using a transgenic approach, we investigated how reducing the extent of O‐acetylation in xylan affects cell wall chemistry, plant performance and the recalcitrance of lignocellulose to saccharification. The Aspergillus niger acetyl xylan esterase AnAXE1 was expressed in Arabidopsis under the control of either the constitutively expressed 35S CAMV promoter or a woody‐tissue‐specific GT43B aspen promoter, and the protein was targeted to the apoplast by its native signal peptide, resulting in elevated acetyl esterase activity in soluble and wall‐bound protein extracts and reduced xylan acetylation. No significant alterations in cell wall composition were observed in the transgenic lines, but their xylans were more easily digested by a β‐1,4‐endoxylanase, and more readily extracted by hot water, acids or alkali. Enzymatic saccharification of lignocellulose after hot water and alkali pretreatments produced up to 20% more reducing sugars in several lines. Fermentation by Trametes versicolor of tissue hydrolysates from the line with a 30% reduction in acetyl content yielded ~70% more ethanol compared with wild type. Plants expressing 35S:AnAXE1 and pGT43B:AnAXE1 developed normally and showed increased resistance to the biotrophic pathogen Hyaloperonospora arabidopsidis, probably due to constitutive activation of defence pathways. However, unintended changes in xyloglucan and pectin acetylation were only observed in 35S:AnAXE1‐expressing plants. This study demonstrates that postsynthetic xylan deacetylation in woody tissues is a promising strategy for optimizing lignocellulosic biomass for biofuel production.     

Characterization of an Acetyl Xylan Esterase from the Anaerobic Fungus Orpinomyces sp. Strain PC-2

A 1,067-bp cDNA, designated axeA, coding for an acetyl xylan esterase (AxeA) was cloned from the anaerobic rumen fungus Orpinomyces sp. strain PC-2. The gene had an open reading frame of 939 bp encoding a polypeptide of 313 amino acid residues with a calculated mass of 34,845 Da. An active esterase using the original start codon of the cDNA was synthesized in Escherichia coli. Two active forms of the esterase were purified from recombinant E. coli cultures. The size difference of 8 amino acids was a result of cleavages at two different sites within the signal peptide. The enzyme released acetate from several acetylated substrates, including acetylated xylan. The activity toward acetylated xylan was tripled in the presence of recombinant xylanase A from the same fungus. Using p-nitrophenyl acetate as a substrate, the enzyme had a K_m of 0.9 mM and a V_max of 785 μmol min⁻¹ mg⁻¹. It had temperature and pH optima of 30°C and 9.0, respectively. AxeA had 56% amino acid identity with BnaA, an acetyl xylan esterase of Neocallimastix patriciarum, but the Orpinomyces AxeA was devoid of a noncatalytic repeated peptide domain (NCRPD) found at the carboxy terminus of the Neocallimastix BnaA. The NCRPD found in many glycosyl hydrolases and esterases of anaerobic fungi has been postulated to function as a docking domain for cellulase-hemicellulase complexes, similar to the dockerin of the cellulosome of Clostridium thermocellum. The difference in domain structures indicated that the two highly similar esterases of Orpinomyces and Neocallimastix may be differently located, the former being a free enzyme and the latter being a component of a cellulase-hemicellulase complex. Sequence data indicate that AxeA and BnaA might represent a new family of hydrolases.     

Xylanase and acetyl xylan esterase activities of XynA,a key subunit of the Clostridium cellulovorans cellulosome for xylan degradation

The Clostridium cellulovorans xynA gene encodes the cellulosomal endo-1,4-beta-xylanase XynA, which consists of a family 11 glycoside hydrolase catalytic domain (CD), a dockerin domain, and a NodB domain. The recombinant acetyl xylan esterase (rNodB) encoded by the NodB domain exhibited broad substrate specificity and released acetate not only from acetylated xylan but also from other acetylated substrates. rNodB acted synergistically with the xylanase CD of XynA for hydrolysis of acetylated xylan. Immunological analyses revealed that XynA corresponds to a major xylanase in the cellulosomal fraction. These results indicate that XynA is a key enzymatic subunit for xylan degradation in C. cellulovorans.     

Purification and characterization of two thermostable acetyl xylan esterases from Thermoanaerobacterium sp. strain JW/SL-YS485.

Two acetyl esterases (EC 3.1.1.6) were purified to gel electrophoretic homogeneity from Thermoanaerobacterium sp. strain JW/SL-YS485, an anaerobic, thermophilic endospore former which is able to utilize various substituted xylans for growth. Both enzymes released acetic acid from chemically acetylated larch xylan. Acetyl xylan esterases I and II had molecular masses of 195 and 106 kDa, respectively, with subunits of 32 kDa (esterase I) and 26 kDa (esterase II). The isoelectric points were 4.2 and 4.3, respectively. As determined by a 2-min assay with 4-methylumbelliferyl acetate as the substrate, the optimal activity of acetyl xylan esterases I and II occurred at pH 7.0 and 80 degrees C and at pH 7.5 and 84 degrees C, respectively. Km values of 0.45 and 0.52 mM 4-methylumbelliferyl acetate were observed for acetyl xylan esterases I and II, respectively. At pH 7.0, the temperatures for the 1-h half-lives for acetyl xylan esterases I and II were 75 degrees and slightly above 100 degrees C, respectively.     

Functional and structural characterization of a thermostable acetyl esterase from Thermotoga maritima

TM0077 from Thermotoga maritima is a member of the carbohydrate esterase family 7 and is active on a variety of acetylated compounds, including cephalosporin C. TM0077 esterase activity is confined to short‐chain acyl esters (C2–C3), and is optimal around 100°C and pH 7.5. The positional specificity of TM0077 was investigated using 4‐nitrophenyl‐β‐D ‐xylopyranoside monoacetates as substrates in a β‐xylosidase‐coupled assay. TM0077 hydrolyzes acetate at positions 2, 3, and 4 with equal efficiency. No activity was detected on xylan or acetylated xylan, which implies that TM0077 is an acetyl esterase and not an acetyl xylan esterase as currently annotated. Selenomethionine‐substituted and native structures of TM0077 were determined at 2.1 and 2.5 Å resolution, respectively, revealing a classic α/β‐hydrolase fold. TM0077 assembles into a doughnut‐shaped hexamer with small tunnels on either side leading to an inner cavity, which contains the six catalytic centers. Structures of TM0077 with covalently bound phenylmethylsulfonyl fluoride and paraoxon were determined to 2.4 and 2.1 Å, respectively, and confirmed that both inhibitors bind covalently to the catalytic serine (Ser188). Upon binding of inhibitor, the catalytic serine adopts an altered conformation, as observed in other esterase and lipases, and supports a previously proposed catalytic mechanism in which Ser hydroxyl rotation prevents reversal of the reaction and allows access of a water molecule for completion of the reaction. Proteins 2012. © 2012 Wiley Periodicals, Inc.     

枯草芽胞杆菌乙酰木聚糖酯酶的鉴定及诱导剂对酯酶活力的影响

以枯草芽胞杆菌CICC 20034为研究对象,对其分泌的高相对分子质量酯酶进行鉴定,并考察诱导剂对其活力的影响。结果表明:枯草芽胞杆菌CICC 20034可分泌一种相对分子质量为1.07×105的酯酶,经蛋白质质谱鉴定为乙酰木聚糖酯酶,单体分相对子质量为3.56×104。在发酵培养基中添加乙酸乙酯和木糖可以显著的促进乙酰木聚糖酯酶的活力,而三丁酸甘油酯和大分子诱导剂——木聚糖、玉米芯粉和壳聚糖对酯酶的活力几乎无促进作用。枯草芽胞杆菌CICC 20034以乙酸乙酯为诱导剂时最高比酶活为0.62 U/mL,为已知报道的野生细菌乙酰木聚糖酯酶的最高酯酶活力。