Putative Ribosome-binding Site Research Articles

Role and Regulation of Poly-3-Hydroxybutyrate in Nitrogen Fixation in Azorhizobium caulinodans

An Azorhizobium caulinodans phaC mutant (OPS0865) unable to make poly-3-hydroxybutyrate (PHB), grows poorly on many carbon sources and cannot fix nitrogen in laboratory culture. However, when inoculated onto its host plant, Sesbania rostrata, the phaC mutant consistently fixed nitrogen. Upon reisolation from S. rostrata root nodules, a suppressor strain (OPS0921) was isolated that has significantly improved growth on a variety of carbon sources and also fixes nitrogen in laboratory culture. The suppressor retains the original mutation and is unable to synthesize PHB. Genome sequencing revealed a suppressor transition mutation, G to A (position 357,354), 13 bases upstream of the ATG start codon of phaR in its putative ribosome binding site (RBS). PhaR is the global regulator of PHB synthesis but also has other roles in regulation within the cell. In comparison with the wild type, translation from the phaR native RBS is increased approximately sixfold in the phaC mutant background, suggesting that the level of PhaR is controlled by PHB. Translation from the phaR mutated RBS (RBS*) of the suppressor mutant strain (OPS0921) is locked at a low basal rate and unaffected by the phaC mutation, suggesting that RBS* renders the level of PhaR insensitive to regulation by PHB. In the original phaC mutant (OPS0865), the lack of nitrogen fixation and poor growth on many carbon sources is likely to be due to increased levels of PhaR causing dysregulation of its complex regulon, because PHB formation, per se, is not required for effective nitrogen fixation in A. caulinodans. [Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY 4.0 International license .

Analysis of Orientia tsutsugamushi promoter activity.

Orientia tsutsugamushi is an obligate intracellular bacterium that causes scrub typhus, a potentially fatal rickettsiosis, and for which no genetic tools exist. Critical to addressing this technical gap is to identify promoters for driving expression of antibiotic resistance and fluorescence reporter genes in O. tsutsugamushi. Such promoters would need to be highly conserved among strains, expressed throughout infection, and exhibit strong activity. We examined the untranslated regions upstream of O. tsutsugamushi genes encoding outer membrane protein A (ompA), 22-kDa type-specific antigen (tsa22) and tsa56. The bacterium transcribed all three during infection of monocytic, endothelial and epithelial cells. Examination of the upstream noncoding regions revealed putative ribosome binding sites, one set of predicted -10 and -35 sequences for ompA and two sets of -10 and -35 sequences for tsa22 and tsa56. Comparison of these regions among geographically diverse O. tsutsugamushi patient isolates revealed nucleotide identities ranging from 84.8 to 100.0%. Upon examination of the candidates for the ability to drive green fluorescence protein expression in Escherichia coli, varying activities were observed with one of the tsa22 promoters being the strongest. Identification and validation of O. tsutsugamushi promoters is an initial key step toward genetically manipulating this important pathogen.

Characterization of a putative ribosome binding site at the 5' untranslated region of bovine heat shock protein 90.

Untranslated regions (UTRs) of the transcripts play significant roles in translation regulation and continue to raise many intriguing questions in our understanding of cellular stress physiology. Internal ribosome entry site (IRES) mediated alternative translation initiations are emerging as unique mechanisms. Present study is aimed to indentify a functional short 92 base pair length putative sequence located at the 5' untranslated region of bovine heat shock protein 90 AA1 (Hsp90AA1) may interact with ribosomal as well as eukaryotic initiation factor binding site. Here we have predicted both the two and three dimensional structures of bovine Hsp90AA1 IRES (MF400854) element with their respective free energy. Molecular interactions between bovine RPS5 and IRES have been determined after the preparation of docking complex of IRES bound RPS5. Structure of bovine ribosomal translational initiation factor (TIF) has also been determined and docked with IRES. Molecular interaction between bovine TIF and IRES was analyzed from the complex structure. We further detected the relative expression efficiency of the viral (original) in relation with Hsp90AA1 IRES-driven GFP expression, which revealed that efficiency under the control of identified bovine Hsp90AA1 IRES was slightly lower than viral origin. It was also noted that identified bovine HSP90 IRES may increase the expression level of GFP under in vitro heat stressed condition.

Reannotation of translational start sites in the genome of Mycobacterium tuberculosis

Identification and correction of incorrect ORF start sites is important for a variety of experimental and analytical purposes, ranging from cloning to inference of operon structure. The genome of the H37Rv reference strain of Mycobacterium tuberculosis (Mtb) was originally annotated when it was first sequenced nearly 15 years ago. While this annotation has served the TB research community well as a standard of reference for over a decade, it has been demonstrated experimentally that the actual start sites for an estimated 5-10% of open reading frames differ from the annotation. In this paper, we present a comprehensive bioinformatic analysis of all 3989 ORFs (open reading frames) in the M. tuberculosis H37Rv genome. Our method combines information from comparative analysis (alignment to start sites of orthologs in other Actinobacteria), sequence conservation, "protein likeness", putative ribosome binding sites, and other data to identify translational start sites. The features are combined in a linear model that is trained on dataset of known start sites verified by mass spectrometry, with a cross-validated accuracy of 94%. The method can be viewed as an augmentation of Hidden Markov Model-based tools such as Glimmer and GeneMark by incorporating more information than just the raw genomic sequence to decide which position is the legitimate translational start site for each ORF. Using this analysis, we identify 269 genes that most likely need to be re-annotated, and identify the best alterative translational start site for each. These revised ORF definitions could be used in the reannotation of the H37Rv genome, as well as to prioritize genes for experimental start-site validation.

Structures of the ribosome-inactivating protein from barley seeds reveal a unique activation mechanism

Ribosome-inactivating protein (RIP), a defence protein found in various plants, possesses different chain architectures and activation mechanisms. The RIP from barley (bRIP) is a type I RIP and has sequence features that are divergent from those of type I and type II RIPs from dicotyledonous plants and even the type III RIP from maize. This study presents the first crystal structure of an RIP from a cereal crop, barley, in free, AMP-bound and adenine-bound states. For phasing, a codon-optimized synthetic brip1 gene was used and a vector was constructed to overexpress soluble bRIP fusion proteins; such expression has been verified in a number of cases. The overall structure of bRIP shows folding similar to that observed in other RIPs but also shows significant differences in specific regions, particularly in a switch region that undergoes a structural transition between a 3(10)-helix and a loop depending on the liganded state. The switch region is in a position equivalent to that of a proteolytically susceptible and putative ribosome-binding site in type III RIPs. Thus, the bRIP structure confirms the detailed enzymatic mechanism of this N-glycosidase and reveals a novel activation mechanism for type I RIPs from cereal crops.

Cloning of transglutaminase gene from Streptomyces platensis YK-2 and its high expression in Streptomyces strains

The microbial transglutaminase (TGase) gene, stgA, was cloned from Streptomyces platensis YK-2, which was newly isolated from a forest soil sample collected in Daegu, Korea. The gene was expressed in several Streptomyces strains including the original strain as the host. The stgA consisted of an open reading frame of 1,257 bp encoding a putative signal peptide of 29 amino acids, a pro-domain of 57 amino acids, and a mature TGase of 332 amino acids, and also had the putative active site of TGase, YGCVG. For expression of the stgA gene, a high expression vector was constructed via the insertion of the entire stgA gene including its putative ribosomal binding site into pSET152ET, located immediately downstream of a strong constitutive ermE* promoter, and was introduced into several Streptomyces strains. The successful high expression and secretion of the correct StgA from streptomycetes were confirmed by electrophoretic sodium dodecyl sulfate polyacrylamide gel electrophoresis and N-terminal amino acid analysis.

A novel cold active esterase derived from Colombian high Andean forest soil metagenome

In order to search new lipolytic enzymes and conduct bioprospecting of microbial communities from high Andean forest soil, a metagenomic library of approximately 20,000 clones was constructed in Escherichia coli using plasmid p-Bluescript II SK+. The library covered 80Mb of the metagenomic DNA mainly from Proteobacteria, Actinobacteria and Acidobacteria. Two clones with lipolytic activity in tributyrin as a substrate were recovered. Clone BAA3G2 (pSK-estGX1) was selected and the entire 4.6Kb insert sequence was determined. The sequence had a GC content of 70.6% and could be derived from an undescribed Actinobacteria genome. One open reading frame encoded a polypeptide of 210 amino acids (gene estGX1) with a molecular mass of 22.4kDa that contained the pentapeptide G-P-S-G-G near the N-terminus essential for lipase activity and the putative catalytic triad was identified, also a putative ribosomal binding site located 18bp upstream the estGX1 ATG start codon was identified. The phylogenetic analysis suggested that the protein belonged to a new lipase family. The secreted enzyme showed a preference for short length fatty acids, with specific activity against p-nitrophenyl-butyrate (0.142U/mg of total protein), it was cold active with relative activity of 30% at 10°C and moderately thermo active with relative activity of 80% at 50°C and had a pH optimum of 8.0 at 40°C.

Searching in microbial genomes for encoded small proteins

As the price of genome sequencing has been rapidly decreasing and can be expected to keep on doing so in the next 10 years, the speed at which new microbial genome sequences become available will increase accordingly. In most genome projects, the first step after acquiring a genome sequence is predicting protein‐encoding open reading frames (pORFs). Small proteins or peptides, loosely defined as less than 50 amino acids, encoded in microbial genomes have been largely underestimated. Recent focused functional genomics efforts have led to the identification of a number of new small proteins encoded in genomes of both Gram‐negative and Gram‐positive bacteria, and fungi (Kastenmayer et al., 2006; Li et al., 2008; Hemm et al., 2010; Hobbs et al., 2010; Bitton et al., 2011). Increasing evidence demonstrates that small proteins participate in a wide array of cellular processes and exhibit great diversity in their mechanisms of action. A recent review (Hobbs et al., 2011) highlights examples of small proteins that, in addition to the well‐conserved small ribosomal proteins, participate in cell signalling or regulation, act as antibiotics and toxins/anti‐toxins, alter membrane features, act as chaperones, stabilize protein complexes or serve as structural proteins (Table 1) (Fig. 1). Table 1 Types and functions of small proteins. Figure 1 Structure of small proteins. Small secreted proteins exhibit diversity in their three‐dimensional structures and can contain unique intramolecular linkages or modified amino acids. For example, the mature form of (A) subtilosin (PDB: 1PXQ) is ... Failure to recognize a pORF encoding a small protein means that these important cell constituents will be missed. Here, we give a brief summary of which problems arise in searching for such encoded small proteins, and what we could do to improve the search process.

Capsule depolymerase overexpression reduces Bacillus anthracis virulence

Capsule depolymerase (CapD) is a gamma-glutamyl transpeptidase and a product of the Bacillus anthracis capsule biosynthesis operon. In this study, we examined the effect of modulating capD expression on B. anthracis capsule phenotype, interaction with phagocytic cells and virulence in guinea pigs. Transcriptional fusions of capD were made to the genes encoding heat-shock protein 60 (hsp60) and elongation factor Tu (EFTu), and to capA, a B. anthracis capsule biosynthesis gene. Translation signals were altered to improve expression of capD, including replacing the putative ribosome-binding site with a consensus sequence and the TTG start codon with ATG. CapD was not detected by immunoblotting in lysates from wild-type B. anthracis Ames but was detected in strains engineered with a consensus ribosome-binding site for capD. Strains overexpressing capD at amounts detected by immunoblotting were found to have less surface-associated capsule and released primarily lower-molecular-mass capsule into culture supernatants. Overexpression of capD increased susceptibility to neutrophil phagocytic killing and adherence to macrophages and resulted in reduced fitness in a guinea pig model of infection. These data suggest that B. anthracis may have evolved weak capD expression resulting in optimized capsule-mediated virulence.

Characterization of a cryptic plasmid pSFKW33 from Shewanella sp. 33B

A cryptic plasmid pSFKW33 from psychrotrophic bacterium Shewanella sp. 33B, an isolate from the Gulf of Gdansk (the Baltic Sea), was sequenced and characterized. It is an 8021 bp circular molecule with 38% GC content, which shows a distinctive nucleotide sequence without homology to other known plasmids. The nucleotide sequence analysis predicts eight open reading frames. The deduced amino acid sequence of ORF-1 shared significant similarity with the phage integrase protein from Pseudomonas putida GB-1. The ORF2 product showed some similarity with the hypothetical protein Shal_1786 from Shewanella halifaxensis HAW-EB4. The ORF-3 product revealed high amino acid sequence homology with the Escherichia coli replication initiation protein similar to pi protein of R6K plasmid. Five repeat regions (three perfect 22-bp repeats and two imperfect motifs), a putative ribosome binding site, and −10 and −35 promotor sequences were identified upstream of the ORF-3 ( rep). The rep module is very similar to several theta replicating iteron family plasmids, suggesting that pSFKW33 replication follows the same course. The ORF-5 product revealed significant identity with the entry exclusion protein 1 of plasmid pIS2 from enteropathogenic strain E. coli 0111:H. The ORF-6 product showed significant identity with the putative transcriptional regulator protein from Shewanella benthica KT99 belonging to HTH-XRE-family protein. The ORF-8 encoded protein that showed some similarity with the hypothetical protein KT99_00146 from Shewanella benthica KT99. The ORF-4 and ORF-7 encoded putative proteins of 189 and 118 amino acid residues, respectively, with no homologies to any known proteins. The absolute copy number of pSFKW33 was estimated to be five copies per chromosome by real-time PCR.

Efficient translation in chloroplasts requires element(s) upstream of the putative ribosome binding site fromatpI

Thousands of proteins make up a chloroplast, but fewer than 100 are encoded by the chloroplast genome. Despite this low number, expression of chloroplast-encoded genes is essential for plant survival. Every chloroplast has its own gene expression system with a major regulatory point at the initiation of protein synthesis (translation). In chloroplasts, most protein-encoding genes contain elements resembling the ribosome binding sites (RBS) found in prokaryotes. In vitro, these putative chloroplast ribosome binding sequences vary in their ability to support translation. Here we report results from an investigation into effects of the predicted RBS for the tobacco chloroplast atpI gene on translation in vivo. Two reporter constructs, differing only in their 5'-untranslated regions (5'UTRs) were stably incorporated into tobacco chloroplast genomes and their expression analyzed. One 5'UTR was derived from the wild-type (WT) atpI gene. The second, Holo-substitution (Holo-sub), had nonchloroplast sequence replacing all wild-type nucleotides, except for the putative RBS. The abundance of reporter RNA was the same for both 5'UTRs. However, translation controlled by Holo-sub was less than 4% that controlled by WT. These in vivo experiments support the idea that translation initiation in land plant chloroplasts depends on 5'UTR elements outside the putative RBS.

Crystal structure of native cinnamomin isoform III and its comparison with other ribosome inactivating proteins

Crystal structure of native cinnamomin isoform III and its comparison with other ribosome inactivating proteins Arezki Azzi,1,2y Tao Wang,1y Dao-Wei Zhu, Yong-Shui Zou, Wang-Yi Liu, and Sheng-Xiang Lin* 1 Structural Biology Platform, Institute of Biochemistry and Cell Biology (IBCB), Shanghai Institutes of Biological Sciences (SIBS), Chinese Academy of Sciences, China 2 Laboratory of Oncology and Molecular Endocrinology, CHUL Research Center (CHUQ) and Laval University, Quebec, Canada

Multiple factors contribute to keeping levels of the symbiosis regulator RscS low

Increased activity alleles (rscS1 and rscS2) of the symbiosis regulator RscS induced both syp transcription and biofilm formation in Vibrio fischeri. Neither allele encodes a protein variant; instead they carry mutations near the putative ribosome-binding site and, in the case of rscS1, an additional silent mutation at codon Leu25. In this study, we found that endogenous levels of RscS are very low under the culture conditions examined and that the increased activity alleles dramatically increased the levels of protein. Of the two mutations present in rscS1, the Leu25 mutation, which replaces a rare with a more common Leu codon, appeared to make the greater contribution to increased activity. Our results suggest that RscS levels are maintained at low levels in the cell by the presence of a weak promoter, possible inefficient ribosome binding and the presence of rare codons in the 5' end of the gene. Restriction of RscS levels may be important to prevent spurious signalling by this sensor kinase in the absence of a squid host.

Isolation and heterologous expression of PHA synthesising genes from Bacillus thuringiensis R1

The polyhydroxyalkanoate biosynthesis gene locus from Bacillus thuringiensis R1 was isolated, cloned and analyzed at the molecular level. We found that a ∼5 kb SacI–ClaI digested fragment of genomic DNA from B. thuringiensis R1 encoding the PHA synthesising genes, conferred PHA producing ability to E. coli. The fragment was sequenced and found to be of 4787 bp with five open reading frames. Sequence alignment with closely related species of Bacillus in the existing database revealed that the ORFs correspond to phaP, phaQ, phaR, phaB and phaC genes. However, E. coli harboring phaP, phaQ, phaR, phaB and phaC locus produced very low PHA. Furthermore, complementation of the locus with phaA from Ralstonia eutropha increased the PHA production in the recombinant E. coli from 3.0% to 24% of cell dry mass. The putative promoter regions and ribosome binding sites were identified for each of the gene. Conserved domains for PHA synthase and aceto-acetyl-coA reductase were also identified. We hence conclude that the PHA operon of Bacillus thuringiensis R1 consists of phaP, phaQ, phaR, phaB, phaC and complementation of the same with phaA is accountable for its high PHA production.

Cloning and Characterization of a Streptomyces Single Module Type Non-ribosomal Peptide Synthetase Catalyzing a Blue Pigment Synthesis

In the present study, we cloned a gene, designated bpsA, which encodes a single module type non-ribosomal peptide synthetase (NRPS) from a D-cycloserine (DCS)-producing Streptomyces lavendulae ATCC11924. A putative oxidation domain is significantly integrated into the adenylation domain of the NRPS, and the condensation domain is absent from the module. When S. lividans was transformed with a plasmid carrying bpsA, the transformed cells produced a blue pigment, suggesting that bpsA is responsible for the blue pigment synthesis. However, to produce the blue pigment in Escherichia coli, the existence of the 4'-phosphopantetheinyl transferase (PPTase) gene from Streptomyces was necessary, in addition to bpsA. The chemical structure of the pigment was determined as 5,5'-diamino-4,4'-dihydroxy-3,3'-diazadiphenoquinone-(2,2'), called indigoidine. The bpsA gene product, designated BPSA, was overproduced in an E. coli host-vector system and purified to homogeneity, demonstrating that the recombinant enzyme prefers L-Gln as a substrate. The in vitro experiment using L-Gln also showed that the blue pigment was formed by the purified BPSA only when the enzyme was phosphopantetheinylated by adding a Streptomyces PPTase purified from E. coli cells. Each site-directed mutagenesis experiment of Lys(598), Tyr(601), Ser(603), and Tyr(608), which are seen in the oxidation domain of BPSA, suggests that these residues are essential for the binding of FMN to the protein and the synthesis of the blue pigment.

Molecular Cloning and Characterization of a Novel β-Agarase, AgaB, from Marine Pseudoalteromonas sp. CY24

Agarases are generally classified into glycoside hydrolase families 16, 50, and 86 and are found to degrade agarose to frequently generate neoagarobiose, neoagarotetraose, or neoagarohexaose as the main products. In this study we have cloned a novel endo-type beta-agarase gene, agaB, from marine Pseudoalteromonas sp. CY24. The novel agarase encoded by agaB gene has no significant sequence similarity with any known proteins including all glycoside hydrolases. It degrades agarose to generate neoagarooctaose and neoagarodecaose as the main end products. Based on the analyses of enzymatic kinetics and degradation patterns of different oligosaccharides, the agarase AgaB appears to have a large substrate binding cleft that accommodates 12 sugar units, with 8 sugar units toward the reducing end spanning subsites +1 to +8 and 4 sugar units toward the non-reducing end spanning subsites -4 to -1, and enzymatic cleavage taking place between subsites -1 and +1. In addition, 1H NMR analysis shows that this enzyme hydrolyzes the glycosidic bond with inversion of anomeric configuration, in contrast to other known agarases that are retaining. Altogether, AgaB is structurally and functionally different from other known agarases and appears to represent a new family of glycoside hydrolase.

Characterization of Strong Promoters from an Environmental Flavobacterium hibernum Strain by Using a Green Fluorescent Protein-Based Reporter System

We developed techniques for the genetic manipulation of Flavobacterium species and used it to characterize several promoters found in these bacteria. Our studies utilized Flavobacterium hibernum strain W22, an environmental strain we isolated from tree hole habitats of mosquito larvae. Plasmids from F. hibernum strain W22 were more efficiently (approximately 1,250-fold) transferred by electroporation into F. hibernum strain W22 than those isolated from Escherichia coli, thus indicating that an efficient restriction barrier exists between these species. The strong promoter, tac, functional in proteobacteria, did not function in Flavobacterium strains. Therefore, a promoter-trap plasmid, pSCH03, containing a promoterless gfpmut3 gene was constructed. A library of 9,000 clones containing chromosomal fragments of F. hibernum strain W22 in pSCH03 was screened for their ability to drive expression of the promoterless gfpmut3 gene. Twenty strong promoters were used for further study. The transcription start points were determined from seven promoter clones by the 5' rapid amplification of cDNA ends technique. Promoter consensus sequences from Flavobacterium were identified as TAnnTTTG and TTG, where n is any nucleotide, centered approximately 7 and 33 bp upstream of the transcription start site, respectively. A putative novel ribosome binding site consensus sequence is proposed as TAAAA by aligning the 20-bp regions upstream of the translational start site in 25 genes. Our primary results demonstrate that at least some promoter and ribosome binding site motifs of Flavobacterium strains are unusual within the bacterial domain and suggest an early evolutionary divergence of this bacterial group. The techniques presented here allow for more detailed genetics-based studies and analyses of Flavobacterium species in the environment.

Constitutive Expression of Outer Surface Protein C Diminishes the Ability ofBorrelia burgdorferiTo Evade Specific Humoral Immunity

The Lyme disease spirochete Borrelia burgdorferi reduces the expression of outer surface protein C (OspC) in response to the development of an anti-OspC humoral response, leading to the hypothesis that the ability to repress OspC expression is critical for the pathogen to proceed to chronic infection. B. burgdorferi was genetically modified to constitutively express OspC by introducing an extra ospC copy fused with the borrelial flagellar gene (flaB) promoter. Such a genetic modification did not reduce infectivity or pathogenicity in severe combined immunodeficiency mice but resulted in clearance of infection by passively transferred OspC antibody. Spirochetes with constitutive ospC expression were unable to establish chronic infections in immunocompetent mice unless they had undergone very destructive mutations in the introduced ospC copy. Two escape mutants were identified; one had all 7 bp deleted between the putative ribosome-binding site and the start codon, ATG, causing a failure in translational initiation, and the other mutant had an insertion of 2 bp between nucleotides 315 and 316, resulting in a nonsense mutation at codon 108. Thus, the ability of B. burgdorferi to repress ospC expression during mammalian infection allows the pathogen to avoid clearance and to preserve the integrity of the important gene for subsequent utilization during its enzootic life cycle.

Cloning and sequencing of plasmid pLC494 isolated from human intestinal Lactobacillus casei: Construction of an Escherichia coli– Lactobacillus shuttle vector

The complete nucleotide sequence of plasmid pLC494 isolated from Lactobacillus casei L-49 was determined. Plasmid pLC494 is an 8846-bp long circular molecule with a G+C content of 41.5%. Two putative open reading frames, ORF4 (282 amino acids) and ORF5 (169 amino acids), were identified as replication proteins A and B that revealed 100 and 99% similarity, respectively, with the replication proteins of plasmid pLA103 from Lactobacillus acidophilus TK8912. Upstream of ORF4 were the four repeat regions (three perfect 22-bp repeats and one imperfect motif), a putative ribosome binding site, a −10 region, and a −35 region. The shuttle vector pJLE4942 (5318 bp) was constructed using repA from pLC494, a multiple cloning site, ColE1 ori, the ori of gram-negative bacteria from vector pUC19, and the chloramphenicol resistance gene from pJIR418 as a selection marker. Transformation of several lactic acid bacteria with the vector pJLE4942 indicated that this vector might be useful as a genetic tool for the intestinal lactobacilli.

Bacillus alcalophilus AX2000 유래 xylanase 유전자 (XynT)의 Cloning과 염기서열 분석

A gene coding for xylanase from alkali-tolerant Bacillus alcalophilus AX2000 was cloned into Escherichia coli using pUC19. Among 2,000 transformants, one transformant showed clear zone on the detection agar plate containing oat-spells xylan. Its recombinant plasmid, named pXTY99, was found to carry 7.0 kb insert DNA fragment. When the nucleotide sequence of the cloned xylanase gene (xynT) was determined, xynT gene was found to consist of 1,020 base-pair open reading frame coding for a poly-peptide of 340 amino acids with a deduced molecular weight of 40 kDa. The coding sequence was preceded by a putative ribosome binding site, and the transcription initiation signals. The deduced amino acid sequence of xylanase is similar to those of the xylanases from Bacillus sp. Nl37 and B. stearothermophilus 21 with and identical residues, respectively