Expression and sequence comparison of the Aspergillus niger and Aspergillus tubigensis genes encoding polygalacturonase II

Bussink, H. J. D.; Buxton, Frank P.; Visser, Jaap

doi:10.1007/bf00312738

Cited by 104 publications

(55 citation statements)

References 45 publications

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“…xyli (Herlache et al, 1997;Huang & Schell, 1990;Hugouvieux-Cotte-Pattat et al, 2002;Liu et al, 1994;Monteiro-Vitorello et al, 2004). Four amino acid groups (NTD, DD, HG and RIK), presumably involved in catalysis (Bussink et al, 1991), are conserved in these polygalacturonases and are situated at aa 243-245, 266-267, 292-293 and 331-333 in Xcc PehA, respectively.…”

Section: Resultsmentioning

confidence: 99%

Regulation of the pehA gene encoding the major polygalacturonase of Xanthomonas campestris by Clp and RpfF

Hsiao

Zheng

et al. 2008

Microbiology

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Exopolysaccharide and several extracellular enzymes of Xanthomonas campestris pv. campestris (Xcc), the causative agent of black rot in crucifers, are virulence factors. In this study, sequence and mutational analysis has demonstrated that Xcc pehA encodes the major polygalacturonase, a member of family 28 of the glycosyl hydrolases. Using the 59 RACE (rapid amplification of cDNA ends) method, the pehA transcription initiation site was mapped at 102 nt downstream of a Clp (cAMP receptor protein-like protein)-binding site. Transcriptional fusion assays showed that pehA transcription is greatly induced by polygalacturonic acid, positively regulated by Clp and RpfF (an enoyl-CoA hydratase homologue which is required for the synthesis of cis-11-methyl-2-dodecenoic acid, a low-molecular-mass diffusible signal factor), subjected to catabolite repression, which is independent of Clp or RpfF, and repressed under conditions of oxygen limitation or nitrogen starvation. Our findings extend previous work on Clp and RpfF regulation to show that they both influence the expression of pehA in Xcc.

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Section: Resultsmentioning

confidence: 99%

Regulation of the pehA gene encoding the major polygalacturonase of Xanthomonas campestris by Clp and RpfF

Hsiao

Zheng

et al. 2008

Microbiology

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“…However, an analysis of a multiple alignment of the enzymes produced different results. Figure 2 shows part of a multiple alignment of four domains of conserved amino acids which were first described for PGs of plant, fungal, and bacterial origin (3). When all of the PGs recovered from a database search were aligned, only the following four short stretches of amino acids were conserved: NXD, DD, HG, and RXK (14) (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…The BCA reagent was freshly prepared by mixing stock solutions A and B (1:1, vol/vol). Solution A contained (per liter of distilled water) 54.3 g of Na 2 CO 3 , 24.2 g of NaHCO 3 , and 1.9 g of Na 2 BCA. Solution B contained (per liter of distilled water) 1.25 g of CuSO 4 ⅐ 5H 2 O and 1.26 g of L-lysine.…”

Section: Methodsmentioning

confidence: 99%

Endo-Xylogalacturonan Hydrolase, a Novel Pectinolytic Enzyme

Vlugt-Bergmans

Meeuwsen²,

Voragen³

et al. 2000

Appl Environ Microbiol

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We screened an Aspergillus tubingensis expression library constructed in the yeast Kluyveromyces lactis for xylogalacturonan-hydrolyzing activity in microwell plates by using a bicinchoninic acid assay. This assay detects reducing carbohydrate groups when they are released from a carbohydrate by enzymatic activity. Two K. lactis recombinants exhibiting xylogalacturonan-hydrolyzing activity were found among the 3,400 colonies tested. The cDNA insert of these recombinants encoded a 406-amino-acid protein, designated XghA, which was encoded by a single-copy gene, xghA. A multiple-sequence alignment revealed that XghA was similar to both polygalacturonases (PGs) and rhamnogalacturonases. A detailed examination of conserved regions in the sequences of these enzymes revealed that XghA resembled PGs more. High-performance liquid chromatography and matrix-assisted laser desorption ionization-time of flight mass spectrometry of the products of degradation of xylogalacturonan and saponified modified hairy regions of apple pectin by XghA demonstrated that this enzyme uses an endo type of mechanism. XghA activity appeared to be specific for a xylose-substituted galacturonic acid backbone.Pectin occurs as constituent of higher-plant cell walls, where it is embedded in the cellulose fibrils. The composition of pectin is different in different plant species and also depends on the age and maturity of the plant part. Most pectin polymers consist of smooth homogalacturonan regions and ramified hairy regions. The smooth regions consist of a linear homogalacturonan backbone. The hairy regions, as identified in apples (28), consist of three different subunits; subunit I is xylogalacturonan (xga) (a galacturonan backbone heavily substituted with xylose), subunit II is a short section of a rhamnogalacturonan backbone that has many relatively long arabinan, galactan, and/or arabinogalactan side chains (the hairs), and subunit III is rhamnogalacturonan composed of alternating rhamnose and galacturonic acid residues.

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“…The role of the peh product in plant tissue maceration along with the other expressed enzymes has been investigated by many researchers [55,56]. The GH-28 polygalacturonases are characterized by the presence of 4 conserved amino acid groups (NTD, DD, HG, and RIK) which were thought to be implicated in the catalytic mechanism [57]. As indicated in Figure 2, the aforementioned conserved amino acid regions were found to be present in the deduced amino acid sequence of the peh encoded protein.…”

Section: Discussionmentioning

confidence: 99%

Molecular Cloning and Expression of Cellulase and Polygalacturonase Genes in E. coli as a Promising Application for Biofuel Production

Ibrahim¹,

Jones²,

Hossenya³

2013

J Pet Environ Biotechnol

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Lignocellulosic biomass has potential for bioethanol, a renewable fuel. A limitation is that bioconversion of the complex lignocellulosic material to simple sugars and then to bioethanol is a challenging process. Recent work has focused on the genetic engineering of a biocatalyst that may play a critical role in biofuel production. Escherichia coli have been considered a convenient host for biocatalysts in biofuel production for its fermentation of glucose into a wide range of short-chain alcohols and production of highly deoxygenated hydrocarbon. The bacterium Pectobacterium carotovorum subsp. carotovorum (P. carotovorum) is notorious for its maceration of the plant cell wall causing soft rot. The ability to destroy plants is due to the expression and secretion of a wide range of hydrolytic enzymes that include cellulases and polygalacturonases. P. carotovorum ATCC™ no. 15359 was used as a source of DNA for the amplification of celB, celC and peh. These genes encode 2 cellulases and a polygalacturonase, respectively. Primers were designed based on published gene sequences and used to amplify the open reading frames from the genomic DNA of P. carotovorum. The individual PCR products were cloned into the pTAC-MAT-2 expression vector and transformed into Escherichia coli. The deduced amino acid sequences of the cloned genes have been analyzed for their catalytically active domains. Estimation of the molecular weights of the expressed proteins was performed using SDS-PAGE analysis and celB, celC and peh products were approximately 29.5 kDa, 40 kDa 41.5 kDa, respectively. Qualitative determination of the cellulase and polygalacturonase activities of the cloned genes was carried out using agar diffusion assays.

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Expression and sequence comparison of the Aspergillus niger and Aspergillus tubigensis genes encoding polygalacturonase II

Cited by 104 publications

References 45 publications

Regulation of the pehA gene encoding the major polygalacturonase of Xanthomonas campestris by Clp and RpfF

Regulation of the pehA gene encoding the major polygalacturonase of Xanthomonas campestris by Clp and RpfF

Endo-Xylogalacturonan Hydrolase, a Novel Pectinolytic Enzyme

Molecular Cloning and Expression of Cellulase and Polygalacturonase Genes in E. coli as a Promising Application for Biofuel Production

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