The <i>abp</i> gene in <i>Geobacillus stearothermophilus</i> T‐6 encodes a GH27 β‐<scp>l</scp>‐arabinopyranosidase

Salama, Rachel; Alalouf, Onit; Tabachnikov, Orly; Zolotnitsky, Gennady; Shoham, G.; Shoham, Yuval

doi:10.1016/j.febslet.2012.05.062

Cited by 16 publications

(28 citation statements)

References 38 publications

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“…As previously suggested (Salama et al, 2012) and further discussed below, Ile67 appeared to be a key residue for the substrate-binding and catalytic specificity of Abp. In order to test this hypothesis, this residue was replaced by aspartic acid, as briefly summarized in the following.…”

Section: Preparation and Characterization Of The Abp-i67d Mutantsupporting

confidence: 60%

“…As mentioned above, Abp is a -l-arabinopyranosidase that removes -l-arabinopyranoside moieties from the side chains of l-arabinose-containing polysaccharides ( Fig. 1; Salama et al, 2012). Sequence-alignment analysis determined that the catalytic residues of Abp are Asp197 and Asp255, with Asp197 serving as the nucleophile and Asp255 as the acid/base for the reaction (Salama et al, 2012).…”

Section: The Active Sitementioning

confidence: 95%

“…-l-arabinopyranose residues from a synthetic substrate (pNP--l-arabinopyranoside) as well as from the native substrates sugar beet arabinan and larch arabinogalactan (Salama et al, 2012).…”

Section: Introductionmentioning

confidence: 99%

“…The Abp monomer is made up of 448 amino acids, with a calculated molecular mass of 51 178 Da. Based on sequence homologies with other GH27 enzymes, it has been suggested that Asp197 is the catalytic nucleophile and Asp255 is the catalytic acid/base (Salama et al, 2012). Recently, we were able to crystallize the wild-type enzyme (Abp-WT) and several of its catalytic mutants, as well as collect full X-ray diffraction data from some of these crystals (Lansky, Salama et al, 2013).…”

Section: Introductionmentioning

confidence: 99%

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Structure–specificity relationships in Abp, a GH27 β-L-arabinopyranosidase fromGeobacillus stearothermophilusT6

Lansky

Salama

Solomon

et al. 2014

Acta Cryst D Biol Crystallogr

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L-Arabinose sugar residues are relatively abundant in plants and are found mainly in arabinan polysaccharides and in other arabinose-containing polysaccharides such as arabinoxylans and pectic arabinogalactans. The majority of the arabinose units in plants are present in the furanose form and only a small fraction of them are present in the pyranose form. The L-arabinan-utilization system in Geobacillus stearothermophilus T6, a Gram-positive thermophilic soil bacterium, has recently been characterized, and one of the key enzymes was found to be an intracellular β-L-arabinopyranosidase (Abp). Abp, a GH27 enzyme, was shown to remove β-L-arabinopyranose residues from synthetic substrates and from the native substrates sugar beet arabinan and larch arabinogalactan. The Abp monomer is made up of 448 amino acids, and based on sequence homology it was suggested that Asp197 is the catalytic nucleophile and Asp255 is the catalytic acid/base. In the current study, the detailed three-dimensional structure of wild-type Abp (at 2.28 Å resolution) and its catalytic mutant Abp-D197A with (at 2.20 Å resolution) and without (at 2.30 Å resolution) a bound L-arabinose product are reported as determined by X-ray crystallography. These structures demonstrate that the three-dimensional structure of the Abp monomer correlates with the general fold observed for GH27 proteins, consisting of two main domains: an N-terminal TIM-barrel domain and a C-terminal all-β domain. The two catalytic residues are located in the TIM-barrel domain, such that their carboxylic functional groups are about 5.9 Å from each other, consistent with a retaining mechanism. An isoleucine residue (Ile67) located at a key position in the active site is shown to play a critical role in the substrate specificity of Abp, providing a structural basis for the high preference of the enzyme towards arabinopyranoside over galactopyranoside substrates. The crystal structure demonstrates that Abp is a tetramer made up of two `open-pincers' dimers, which clamp around each other to form a central cavity. The four active sites of the Abp tetramer are situated on the inner surface of this cavity, all opening into the central space of the cavity. The biological relevance of this tetrameric structure is supported by independent results obtained from size-exclusion chromatography (SEC), dynamic light-scattering (DLS) and small-angle X-ray scattering (SAXS) experiments. These data and their comparison to the structural data of related GH27 enzymes are used for a more general discussion concerning structure-selectivity aspects in this glycoside hydrolase (GH) family.

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Section: Preparation and Characterization Of The Abp-i67d Mutantsupporting

confidence: 60%

Section: The Active Sitementioning

confidence: 95%

“…-l-arabinopyranose residues from a synthetic substrate (pNP--l-arabinopyranoside) as well as from the native substrates sugar beet arabinan and larch arabinogalactan (Salama et al, 2012).…”

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Structure–specificity relationships in Abp, a GH27 β-L-arabinopyranosidase fromGeobacillus stearothermophilusT6

Lansky

Salama

Solomon

et al. 2014

Acta Cryst D Biol Crystallogr

Self Cite

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“…Thermophilic endospore-formers constitute an excellent resource of biotechnologically important products and can be used as the producers of these proteins [6][7][8][9][10]. It is essential to monitor the purity of the protein-producing strains in these processes.…”

Section: Introductionmentioning

confidence: 99%

Characterization and evaluation of tandem repeats for the identification of Geobacillus

Krilavičiūtė

Kuisienė

2013

Open Life Sciences

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Taxonomy of thermophilic, endospore-forming bacteria has evoked a great interest over the past few years. Although a number of taxonomic markers were previously evaluated, their sequences in Geobacillus were too conservative, and identification of more variable markers is needed. Repetitive DNA is one of the promising variable targets in the development of the taxon-specific genotyping and identification schemes in bacteria. The aim of our study was to evaluate the possibility of using repetitive DNA in the taxonomy of Geobacillus. In this paper, we report the analysis of perfect tandem repeats of geobacilli. We focused on the long repeats (with a motif length of ≥20 nucleotides). This choice was based on the assumption that these motifs can be used for the construction of oligonucleotides — primers and probes. Thirty-three Geobacillus genus-specific motifs were identified in our work, fifteen of them were species-specific and fifteen — species cluster -specific. Three of them were genus-, but not species- or species cluster-specific. Some of the motifs were used for the construction of the primer pairs. The primers were validated by PCR. Out of 12 designed primer pairs, 11 were genus-specific and 4 — species-specific. Species-specific primers were successfully constructed for the phylogenetically defined species Geobacillus thermodenitrificans and Geobacillus toebii.

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Metabolism of l-arabinose in plants

et al. 2016

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l-Arabinose (l-Ara) is a plant-specific sugar accounting for 5–10 % of cell wall saccharides in Arabidopsis (Arabidopsis thaliana) and rice (Oryza sativa). l-Ara occurs in pectic arabinan, rhamnogalacturonan II, arabinoxylan, arabinogalactan-protein (AGP), and extensin in the cell walls, as well as in glycosylated signaling peptides like CLAVATA3 and small glycoconjugates such as quercetin 3-O-arabinoside. This review focuses on recent advances towards understanding the generation of l-Ara and the metabolism of l-Ara-containing molecules in plants.Electronic supplementary materialThe online version of this article (doi:10.1007/s10265-016-0834-z) contains supplementary material, which is available to authorized users.

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The abp gene in Geobacillus stearothermophilus T‐6 encodes a GH27 β‐l‐arabinopyranosidase

Cited by 16 publications

References 38 publications

Structure–specificity relationships in Abp, a GH27 β-L-arabinopyranosidase fromGeobacillus stearothermophilusT6

Structure–specificity relationships in Abp, a GH27 β-L-arabinopyranosidase fromGeobacillus stearothermophilusT6

Characterization and evaluation of tandem repeats for the identification of Geobacillus

Metabolism of l-arabinose in plants

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