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At least 811 records · Page 45Linked to original sources
PubMed2006

Amino acid regions of family 45 endoglucanases involved in cotton defibrillation and in resistance to anionic surfactants and oxidizing agents.

In the detergent industry, fungal endoglucanases are used to release microfibrils from the surfaces of dyed cellulosic fabrics to enhance color brightness. Family 45 endoglucanase (glycoside hydrolase family 45, GH45) EGL3 from Humicola grisea is more resistant to anionic surfactants and oxidizing agents than family 45 endoglucanase RCE1 from Rhizopus oryzae, while in the present study, a catalytic domain of RCE1 had higher defibrillation activity on dyed cotton fabrics than did that of EGL3. To identify the amino acid regions involved in these properties, we compared the characteristics of RCE1, EGL3, and three chimeric endoglucanases, in which each of the three regions of the catalytic domain of EGL3 was replaced by the corresponding region of the catalytic domain of RCE1. Amino acids in the N-terminal region were involved in resistance to anionic surfactants and oxidizing agents. Furthermore, amino acids in the region adjacent to the N-terminal region were involved in releasing microfibrils and in binding to dyed cotton fabrics, indicating that the binding of the amino acids in this region might be important in the release of microfibrils from dyed cotton fabrics.

Amino Acids↗
PubMed2000

Purification and characterization of an alpha-L-rhamnosidase from Pichia angusta X349.

An intracellular alpha-L-rhamnosidase from Pichia angusta X349 was purified to homogeneity through four chromatographic steps. The alpha-L-rhamnosidase appeared to be a monomeric protein with a molecular mass of 90 kDa. The enzyme had an isoelectric point at 4.9, and was optimally active at pH 6.0 and at around 40 degrees C. The Ki for L-rhamnose inhibition was 25 mM. The enzyme was inhibited by Cu2+, Hg2+, and p-chloromercuribenzoate. The alpha-L-rhamnosidase was highly specific for alpha-L-rhamnopyranoside and liberated rhamnose from naringin, rutin, hesperidin, and 3-quercitrin. The alpha-L-rhamnosidase was active at the ethanol concentrations of wine. It efficiently released monoterpenols, such as linalool and geraniol, from an aroma precursor extracted from Muscat grape juice.

Amino Acid Sequence↗
PubMed2000

Purification and characterization of a family G/11 beta-xylanase from Streptomyces olivaceoviridis E-86.

A beta-xylanase (GXYN) was purified from the culture filtrate of Streptomyces olivaceoviridis E-86 by successive chromatography on DE-52, CM-Sepharose and Superose 12. The molecular mass of the xylanase was estimated to be 23 kDa, indicating that the enzyme consists of a catalytic domain only. The enzyme displayed an optimum pH of 6, a temperature optimum of 60 degrees C, a pH stability range from 2 to 11 and thermal stability up to 40 degrees C. The N-terminal amino acid sequence of GXYN was A-T-V-I-T-T-N-Q-T-G-T-N-N-G-I-Y-Y-S-F-W-, and sharing a high degree of similarity with the N-terminal sequence of xylanases B and C from Streptomyces lividans, indicating GXYN belongs to family G/11 of glycoside hydrolases. GXYN was inferior to xylanase B from Streptomyces lividans in the hydrolysis of insoluble xylan because of its lack of a xylan binding domain.

Amino Acid Sequence↗
PubMed2002

Cloning and sequencing of the genes encoding cyclic tetrasaccharide-synthesizing enzymes from Bacillus globisporus C11.

The genes for isomaltosyltransferase (CtsY) and 6-glucosyltransferase (CtsZ), involved in synthesis of a cyclic tetrasaccharide from alpha-glucan, have been cloned from the genome of Bacillus globisporus C11. The amino-acid sequence deduced from the ctsY gene is composed of 1093 residues having a signal sequence of 29 residues in its N-terminus. The ctsZ gene encodes a protein consisting of 1284 residues with a signal sequence of 35 residues. Both of the gene products show similarities to alpha-glucosidases belonging to glycoside hydrolase family 31 and conserve two aspartic acids corresponding to the putative catalytic residues of these enzymes. The two genes are linked together, forming ctsYZ. The DNA sequence of 16,515 bp analyzed in this study contains four open reading frames (ORFs) upstream of ctsYZ and one ORF downstream. The first six ORFs, including ctsYZ, form a gene cluster, ctsUVWXYZ. The amino-acid sequences deduced from ctsUV are similar in to a sequence permease and a sugar-binding protein for the sugar transport system from Thermococcus sp. B1001. The third ctsW encodes a protein similar to CtsY, suggested to be another isomaltosyltransferase preferring panose to high-molecular-mass substrates.

Amino Acid Sequence↗
PubMed2003

Cloning, sequencing, and expression of the gene encoding the Clostridium stercorarium alpha-galactosidase Aga36A in Escherichia coli.

The alpha-galactosidase gene aga36A of Clostridium stercorarium F-9 was cloned, sequenced, and expressed in Escherichia coli. The aga36A gene consists of 2,208 nucleotides encoding a protein of 736 amino acids with a predicted molecular weight of 84,786. Aga36A is an enzyme classified in family 36 of the glycoside hydrolases and showed sequence similarity with some enzymes of family 36 such as Geobacillus (formerly Bacillus) stearothermophilus GalA (57%) and AgaN (52%). The enzyme purified from a recombinant E. coli is optimally active at 70 degrees C and pH 6.0. The enzyme hydrolyzed raffinose and guar gum with specific activities of 3.0 U/mg and 0.46 U/mg for the respective substrates.

Bacterial Proteins↗
PubMed2004

Cloning of the Ruminococcus albus cel5D and cel9A genes encoding dockerin module-containing endoglucanases and expression of cel5D in Escherichia coli.

An EcoRI chromosomal DNA fragment of Ruminococcus albus F-40 that conferred endoglucanase activity on Escherichia coli was cloned. An open reading frame (ORF1) and another incomplete reading frame (ORF2) were found in the EcoRI fragment. The ORF2 was completed using inverse PCR genome walking technique. ORF1 and ORF2, which confront each other, encoded cellulases belonging to families 5 and 9 of the glycoside hydrolases and were designated cel5D and cel9A respectively. The cel5D gene encodes 753 amino acids with a deduced molecular weight of 83,409. Cel5D consists of a signal peptide of 24 amino acids, a family-5 catalytic module, a dockerin module, and two family-4 carbohydrate-binding modules (CBMs). The cel9A gene encodes 936 amino acids with a deduced molecular weight of 104,174, consisting of a signal peptide, a family-9 catalytic module, a family-3 CBM, and a dockerin module. The catalytic module polypeptide (rCel5DCat) derived from Cel5D was constructed, expressed, and purified from a recombinant E. coli. The truncated enzyme hydrolyzed cellohexaose, cellopentaose, and cellotetraose to yield mainly cellotriose and cellobiose with glucose as a minor product, but the enzyme was less active toward cellotriose and not active toward cellobiose, suggesting that this enzyme is a typical endoglucanase. rCel5DCat had a Km of 3.9 mg/ml and a Vmax of 37.2 micromol/min/mg for carboxymethycellulose.

Amino Acid Sequence↗
PubMed2004

Molecular cloning of a gene encoding endo-beta-D-1,4-glucanase PCE1 from Phycomyces nitens.

We previously cloned three endoglucanase genes, rce1, rce2, and rce3, from Rhizopus oryzae as the first cellulase genes from the subdivision Zygomycota. In this study, an endoglucanase gene, designated a pce1 gene, was cloned by plaque hybridization with the codon usage-optimized rce1 gene as a probe from Phycomyces nitens, a member of the subdivision Zygomycota. The pec1 gene had an open reading frame of 1,038 nucleotides encoding an endoglucanase (PCE1) of 346 amino acid residues. The amino acid sequence deduced from the pce1 gene consisted of a cellulose-binding domain (CBD) at the N terminus and of a catalytic domain belonging to family 45 glycoside hydrolase at the C terminus. PCE1 was purified to apparent homogeneity from the culture supernatant of P. nitens and the molecular mass was found to be 45 kDa. The optimum pH for the CMCase activity of PCE1 was 6.0, and the optimum temperature was 50 degrees C, the lowest among the family 45 endoglucanases.

Amino Acid Sequence↗
PubMed2004

Cyclic tetrasaccharide-synthesizing enzymes from Arthrobacter globiformis A19.

A bacterial strain Arthrobacter globiformis A19 producing cyclic tetrasaccharide (CTS) was isolated from soil. The enzymes, 6-alpha-glucosyltransferase (6GT) and 3-alpha-isomaltosyltransferase (IMT), involved in the synthesis of CTS were purified to homogeneity. The molecular and enzymatic properties of IMT from A. globiformis were similar to those of enzymes from Bacillus globisporus C11 and N75. Arthrobacter 6GT had a smaller molecular mass of 108 kDa and a higher optimum pH of 8.4 than the enzymes from strains of B. globisporus. The genes for IMT (ctsY) and 6GT (ctsZ) were cloned from the genome of A. globiformis A19. The two genes linked together in tandem and formed a gene cluster, ctsYZ. Both of the gene products showed similarities to alpha-glucosidases belonging to glycoside hydrolase family 31, and conserved two aspartic acids corresponding to the putative catalytic residues of the family enzymes. The enzymatic system for the production of CTS consisting of 6GT and IMT might be widespread among bacteria.

Amino Acid Sequence↗
PubMed2004

Sequencing and expression of the gene encoding the Clostridium stercorarium beta-xylosidase Xyl43B in Escherichia coli.

The Clostridium stercorarium F-9 xyl43B gene encoding the beta-xylosidase Xyl43B consists of an open reading frame of 1,491 nucleotides that encodes a putative protein, classified in family 43, of 497 amino acids with a predicted molecular weight of 56,355. The deduced amino acid sequence of Xyl43B has sequence similarity with beta-xylosidases from Bacteriodes thetaiotaomicron (57% sequence identity), Prevotella ruminicola (45%), Streptomyces coelicolor (40%), and Clostridium acetobutylicum (36%), all of which have been classified in family 43 of the glycoside hydrolases. Xyl43B was purified from a recombinant Escherichia coli and characterized. The optimum pH of the purified enzyme was 3.5 and it was stable over pH from 3.0 to 8.0. Its optimum temperature was 80 degrees C and it showed thermostability in the temperature range from 50 to 70 degrees C. Xyl43B had a K(m) of 6.2 mM and a V(max) of 15 micromol min(-1) mg(-1) for p-nitrophenyl-beta-D-xylopyranoside.

Amino Acid Sequence↗
PubMed2005

Purification and characterization of a new endo-1,4-beta-D-glucanase from Beltraniella portoricensis.

A new endoglucanase, designated BCE1, produced by Beltraniella portoricensis, was purified from the culture supernatant. The N-terminal amino acid sequence suggests that BCE1 belongs to family 45 glycoside hydrolase (family 45 endoglucanase). The molecular mass of BCE1 was found to be 40 kDa by sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE). The optimum pH for the carboxymethyl cellulase (CMCase) activity of BCE1 was 4.5, and the optimum temperature was 55 degrees C. Among family 45 endoglucanases, RCE1 and RCE2 from Rhizopus oryzae, PCE1 from Phycomyces nitens, and EGL3 and EGL4 from Humicola grisea, BCE1 was most resistant to anionic surfactant and oxidizing agent. These results indicate that BCE1 might prove to be a useful enzyme in the detergent industry.

Amino Acid Sequence↗
PubMed2005

Functions of family-22 carbohydrate-binding module in Clostridium thermocellum Xyn10C.

Clostridium thermocellum xylanase Xyn10C (formerly XynC) is a modular enzyme, comprising a family-22 carbohydrate-binding module (CBM), a family-10 catalytic module of the glycoside hydrolases, and a dockerin module responsible for cellulosome assembly consecutively from the N-terminus. To study the functions of the CBM, truncated derivatives of Xyn10C were constructed: a recombinant catalytic module polypeptide (rCM), a family-22 CBM polypeptide (rCBM), and a polypeptide composed of the family-22 CBM and CM (rCBM-CM). The recombinant proteins were characterized by enzyme and binding assays. Although the catalytic activity of rCBM-CM toward insoluble xylan was four times higher than that of rCM toward the same substrate, removal of the CBM did not severely affect catalytic activity toward soluble xylan or beta-1,3-1,4-glucan. rCBM showed an affinity for amorphous celluloses and insoluble and soluble xylan in qualitative binding assays. The optimum temperature of rCBM-CM was 80 degrees C and that of rCM was 60 degrees C. These results indicate that the family-22 CBM of C. thermocellum Xyn10C not only was responsible for the binding of the enzyme to the substrates, but also contributes to the stability of the CM in the presence of the substrate at high temperatures.

Base Sequence↗
PubMed2006

Specific characteristics of family 45 endoglucanases from Mucorales in the use of textiles and laundry.

We examined the characteristics of family 45 endoglucanases (glycoside hydrolases family 45; GH45) from Mucorales belonging to Zygomycota in the use of textiles and laundry. The defibrillation activities on lyocell fabric of family 45 endoglucanases from Mucorales, such as RCE1 and RCE2 from Rhizopus oryzae, MCE1 and MCE2 from Mucor circinelloides, and PCE1 from Phycomyces nitens, were much higher than those of the other family 45 endoglucanases. By contrast, family 45 endoglucanases from Mucorales were less resistant to anionic surfactant and oxidizing agent, main components in detergents, than the other family 45 endoglucanases. RCE1 consists of two distinct modules, a catalytic module and a carbohydrate-binding module family 1 (CBM1), and these common specific characteristics were considered to due to the catalytic module, but not to the CBM1.

Cellulase↗
PubMed2006

Identification of a catalytic residue of Clostridium paraputrificum N-acetyl-beta-D-glucosaminidase Nag3A by site-directed mutagenesis.

Clostridium paraputrificum M-21 beta-N-acetylglucosaminidase 3A (Nag3A) is an enzyme classified in family 3 of the glycoside hydrolases. To identify catalytic residues of this enzyme, mutations were introduced into highly conserved Glu and Asp residues. Replacement of Asp175 with Ala abolished the catalytic activity without change in the circular dichroism spectrum, strongly suggesting that this residue is a catalytic residue, a nucleophile/base or a proton donor. Since the K(m) values of mutant enzymes D119N, D229N, D229A and D274N increased 17 to 41 times as compared with that of wild-type enzyme, Asp119, Asp229, and Asp274 appear to be involved in substrate recognition and binding. Taking previous studies into consideration, we presume that Asp303 is the catalytic nucleophile and Asp175 is the proton donor of C. paraputrificum Nag3A.

Acetylglucosaminidase↗
PubMed2006

Isolation and characterization of a beta-primeverosidase-like enzyme from Penicillium multicolor.

p-Nitrophenyl and eugenyl beta-primeveroside (6-O-beta-D-xylopyranosyl-beta-D-glucopyranoside) hydrolytic activity was found in culture filtrate from Penicillium multicolor IAM7153, and the enzyme was isolated. The enzyme was purified as a beta-primeverosidase-like enzyme by precipitation with ammonium sulfate followed by successive chromatographies on Phenyl Sepharose, Mono Q, and beta-galactosylamidine affinity columns. The molecular mass was estimated to be 50 kDa by SDS-PAGE and gel filtration. The purified enzyme was highly specific toward the substrate p-nitrophenyl beta-primeveroside, which was cleaved in an endo-manner into primeverose and p-nitrophenol, but a series of beta-primeveroside as aroma precursors were hydrolyzed only slightly as substrates for the enzyme. In analyses of its hydrolytic action and kinetics, the enzyme showed narrow substrate specificity with respect to the aglycon and glycon moieties of the diglycoside. We conclude that the present enzyme is a kind of beta-diglycosidase rather than beta-primeverosidase.

Chromatography, Affinity↗
PubMed2024

Characterizing the gut microbiome of diarrheal mink under farmed conditions: A metagenomic analysis.

This study aimed to comprehensively characterize the gut microbiota in diarrheal mink. We conducted Shotgun metagenomic sequencing on samples from five groups of diarrheal mink and five groups of healthy mink. The microbiota α-diversity and Kyoto Encyclopedia of Genes and Genomes (KEGG) orthology did not show significant differences between the groups. However, significant differences were observed in microbiota β-diversity and the function of carbohydrate-active enzymes (CAZymes) between diarrheal and healthy mink. Specifically, The relative abundance of Firmicutes was lower, whereas that of Bacteroidetes was higher in diarrheal mink. Fusobacteria were enriched as invasive bacteria in the gut of diarrheal mink compared with healthy mink. In addition, Escherichia albertii was identified as a new bacterium in diarrheal mink. Regarding functions, nicotinate and nicotinamide metabolism and glycoside hydrolases 2 (GH2) family were the enhanced KEGG orthology and CAZymes in diarrheal mink. Furthermore, the diversity and number of antibiotic-resistant genes were significantly higher in the diarrheal mink group than in the healthy group. These findings enhance our understanding of the gut microbiota of adult mink and may lead to new approaches to the diagnosis and treatment of mink diarrhea.

Animals↗
PubMedDate not supplied

A study on tea aroma formation mechanism: alcoholic aroma precursor amounts and glycosidase activity in parts of the tea plant.

We have shown in molecular basis that alcoholic tea aroma is mainly formed by endogenous enzymatic hydrolysis of glycosidic aroma precursors during manufacturing. Amounts of alcoholic aroma precursor and glycosidase activity in each part of the tea shoot (Camellia sinensis var. sinensis cv Yabukita and a hybrid of var. assamica & var. sinensis cv Izumi) were indirectly measured by means of a crude enzyme assay. The aroma precursors were abundant in young leaves and decreased as the leaf aged. Glycosidase activity also decreased as leaves aged, but was high in stems.

Carbohydrate Conformation↗
PubMed2004

The role of cis-acting sequences governing catabolite repression control of lacS expression in the archaeon Sulfolobus solfataricus.

The archaeon Sulfolobus solfataricus uses a catabolite repression-like system to control production of several glycoside hydrolases. To better understand this regulatory system, studies of the regulation of expression of the beta-glycosidase gene (lacS) were conducted. Expression of lacS varies in response to medium composition and to mutations at an unlinked gene called car. Despite gene overlap, expression of the lacS promoter proximal gene, SSO3017, exhibited coregulation but not cotranscription with lacS. Measurements of mRNA half-life excluded differential stability as a factor in lacS regulation. Chromosomal repositioning by homologous recombination of a lacS deletion series clarified critical cis-acting sequences required for lacS regulation. lacS repositioned at amyA exhibited increased lacS expression and compromised the response to medium composition independently of lacS 5' flanking sequence composition. In contrast, regulation of lacS by the car mutation was dependent on sequences upstream of the archaeal TATA box. Expression of a promoter fusion between lacS and the car-independent malA promoter integrated either at amyA or at the natural lacS locus was insensitive to the allelic state of car. In contrast, the promoter fusion retained a response to medium composition only at the lacS locus. These results indicate that car acts at the lacS promoter and that the response to medium composition involves locus-specific sequences exclusive of those present 5' to lacS or within the lacS transcription unit.

Archaeal Proteins↗
PubMed2004

Triplets! Unexpected structural similarity among the three enzymes that catalyze initiation and termination of thyroid hormone effects.

The three iodothyronine deiodinases catalyze the initiation (D1, D2) and termination (D3) of thyroid hormone effects in vertebrates. A recently conceived 3-dimensional model predicts that these enzymes share a similar structural organization and belong to the thioredoxin (TRX) fold superfamily. Their active center is a selenocysteine-containing pocket defined by the beta1-alpha1-beta2 motifs of the TRX fold and a domain that shares strong similarities with the active site of iduronidase, a member of the clan GH-A fold of glycoside hydrolases. While D1 and D3 are long-lived plasma membrane proteins, D2 is an endoplasmic reticulum resident protein with a half-life of only 20 min. D2 inactivation is mediated by selective UBC-7-mediated conjugation to ubiquitin, a process that is accelerated by T4 catalysis, thus maintaining local T3 homeostasis. In addition, D2 interacts with and is a substrate of the pVHL-interacting deubiquitinating enzymes (VDU1 and VDU2); thus deubiquitination regulates the supply of active thyroid hormone in D2-expressing cells.

Animals↗