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At least 1,567 records · Page 87Linked to original sources
PubMed2006

Leaf hairs influence phytopathogenic fungus infection and confer an increased resistance when expressing a Trichoderma alpha-1,3-glucanase.

The leaf surface of a very large number of plant species are covered by trichomes. Non-glandular trichomes are specialized unicellular or multicellular structures that occur in many different plant species and function in xenobiotic detoxification and protecting the plant against pest attack. By analysing the susceptibility of trichome mutants, evidence is provided that indicates the influence of leaf trichomes on foliar fungal infections in Arabidopsis thaliana, probably by facilitating the adhesion of the fungal spores/hyphae to the leaf surface. A decreased trichome number in the hairless Arabidopsis mutant gl1 enhances tolerance against the necrotrophic fungus Botrytis cinerea. By contrast, the try mutant shows an increased susceptibility to both fungal infection and accumulation. Trichome density does not influence infection by the soil-borne pathogen Rhizoctonia solani. In addition, the influence of trichomes on foliar infection is supported by targeting the high-level expression of the Trichoderma harzianum alpha-1,3-glucanase protein to the specialized cell structures. Trichome expression of this anti-fungal hydrolase shows a significant resistance to infection by the foliar pathogen Botrytis cinerea. Resistance to this fungus is not dependent on the constitutive induction of the salicylic or jasmonic defence signalling pathways, but the presence of the alpha-1,3-glucanase protein in trichomes.

Arabidopsis↗
PubMed1988

The structure of fucosylated blood group substances in fetal rat skin. The combined use of monoclonal antibodies and glycosyl hydrolases.

Cell surface carbohydrates are thought to be important during embryonic development. We have studied a group of defined cell surface carbohydrates during the differentiation of fetal rat skin by the use of immunocytochemical techniques. We used monoclonal antibodies to the LeX and human blood group H type II haptens. The structure of the molecules carrying these haptens has been inferred by the sensitivity of the immune reactions to digestion by glycosyl hydrolases. The primitive ectoderm expressed the LeX hapten. The differentiating epidermis carries the H type II antigen, but this was only detected following cleavage of a terminally modified portion of the carbohydrate containing linear poly-N-acetyl lactosamine chains. These carbohydrates on epidermal cells are carried N linked to glycoproteins.

Animals↗
PubMed2001

Thioglucosidase activity from Sphingobacterium sp. strain OTG1.

Screening for novel thioglucoside hydrolase activity resulted in the isolation of Sphingobacterium sp. strain OTG1 from enrichment cultures containing octylthioglucoside (OTG). OTG was hydrolysed into octanethiol and glucose by cell free extracts. Besides thioglucoside hydrolysis, several other glucoside hydrolase activities were detected in the Sphingobacterium sp. strain OTG1 cell free extract. By adding beta-glucosidase inhibitors it was possible to discriminate between these different activities. Ascorbic acid and D-gluconic acid lactone inhibited the hydrolysis of p-nitrophenyl beta-glucoside, but did not affect octyl- and octylthioglucoside hydrolase activity. Besides OTG, various other thioglucosides were hydrolysed by the novel thioglucosidase, with almost the same activities regardless of the nature of the aglycone, including the myrosinase model substrate sinigrin (a glucosinolate). Sinigrin could also be used as a growth substrate by Sphingobacterium sp. strain OTG1, although at concentrations exceeding 0.15 mM degradation was not complete.

Bacteria↗
PubMed1989

Selective presence of acid hydrolases in the interphotoreceptor matrix.

Adler and Martin (1983, Curr. Eye Res. 2, 359-66) found cathepsin D to be present in crude preparations of bovine interphotoreceptor matrix (IPM). The purpose of the present study was to determine, by investigating several acid hydrolases in purer IPM samples, whether hydrolytic enzymes abundant in RPE lysosomes were present also as normal components of the IPM. IPM was prepared from bovine eyes by the introduction of a small bleb of buffer between the neural retina and the RPE. These IPM samples were free from significant contamination by surrounding tissues; they contained IRBP as their only major protein, and had negligible amounts of lactate dehydrogenase and ROS-specific proteins. Most acid hydrolases were assayed fluorometrically by measuring the 4-methylumbelliferone released upon hydrolysis of appropriate derivatives; the substrate for cathepsin was hemoglobin. The amounts of the enzymes found in the IPM were far from uniform and could not be correlated with enzyme activities in either RPE or retina homogenates. The hydrolases in the IPM varied in amount from beta-galactosidase (28% of the RPE level), through N-acetyl-beta-glucosaminidase (20%), alpha-fucosidase (15%), beta-glucuronidase (12%), alpha-glucosidase (8%), cathepsin D (7%), alpha-mannosidase (7%), down to beta-glucosidase, acid phosphatase, and acid lipase (trace amounts, less than 1%). These results agree with the relative amounts of enzymes found by Wilcox (1987) to be secreted into the medium by cultured human RPE cells. Furthermore, the rank order of hydrolases in the IPM is the same as that for hydrolases secreted (but not recaptured) by human fibroblasts in I-cell disease. The conclusion from these correlations is that lysosomal enzymes are probably secreted, as a normal process, by the RPE into the IPM, where they may have a role in digesting shed outer segments and in catabolizing IPM components.

Acid Phosphatase↗
PubMed1997

Cloning and characterization of two rhamnogalacturonan hydrolase genes from Aspergillus niger.

A rhamnogalacturonan hydrolase gene of Aspergillus aculeatus was used as a probe for the cloning of two rhamnogalacturonan hydrolase genes of Aspergillus niger. The corresponding proteins, rhamnogalacturonan hydrolases A and B, are 78 and 72% identical, respectively, with the A. aculeatus enzyme. In A. niger cultures which were shifted from growth on sucrose to growth on apple pectin as a carbon source, the expression of the rhamnogalacturonan hydrolase A gene (rhgA) was transiently induced after 3 h of growth on apple pectin. The rhamnogalacturonan hydrolase B gene was not induced by apple pectin, but the rhgB gene was derepressed after 18 h of growth on either apple pectin or sucrose. Gene fusions of the A. niger rhgA and rhgB coding regions with the strong and inducible Aspergillus awamori exlA promoter were used to obtain high-producing A. awamori transformants which were then used for the purification of the two A. niger rhamnogalacturonan hydrolases. High-performance anion-exchange chromatography of oligomeric degradation products showed that optimal degradation of an isolated highly branched pectin fraction by A. niger rhamnogalacturonan hydrolases A and B occurred at pH 3.6 and 4.1, respectively. The specific activities of rhamnogalacturonan hydrolases A and B were then 0.9 and 0.4 U/mg, respectively, which is significantly lower than the specific activity of A. aculeatus rhamnogalacturonan hydrolase (2.5 U/mg at an optimal pH of 4.5). Compared to the A enzymes, the A. niger B enzyme appears to have a different substrate specificity, since additional oligomers are formed.

Amino Acid Sequence↗
PubMed1971

Studies on the structure-bound sedimentabolity of some rat liver lysosome hydrolases.

1. Lysosome-rich fractions from rat liver were subjected to several disruptive procedures: osmotic lysis or freezing and thawing in different media, shearing forces in a high-speed blender, treatment with Triton X-100. 2. The soluble and particulate phases were then separated by high-speed centrifugation and assayed for their content of acid phosphatase, beta-galactosidase, beta-N-acetylglucosaminidase, acid proteinase, acid ribonuclease, acid deoxyribonuclease and protein. 3. The degree of elution of these hydrolases appeared to depend on both the enzyme species and the treatment. The resulting patterns of solubilization were rather complex, so that a clear-cut discrimination between soluble and structure-bound enzymes could not always be traced. 4. Although only beta-galactosidase was readily solubilizable after all treatments, acid proteinase could also be extensively eluted from the sedimentable material in the presence of EDTA and acid phosphatase was fully extracted by Triton X-100. On the other hand, considerable proportions of the other activities could not be solubilized by any of the procedures used. 5. In other experiments, the adsorbability of hydrolases on subcellular structures was investigated by measuring the partition between sedimentable particles and soluble fraction of solubilized enzymes added to ;intact' liver homogenates. 6. Large proportions of acid proteinase, ribonuclease and deoxyribonuclease, and almost all of beta-N-acetylglucosaminidase, were found to be adsorbed on the particulate material.

Acid Phosphatase↗
PubMed1984

In vivo and in vitro evidence for the involvement of cysteine proteinases in bone resorption.

The excretion of cathepsin B, a lysosomal cysteine proteinase, by parathyroid hormone-stimulated embryonic mouse calvaria in culture, correlates closely with the extent of bone resorption evaluated by the loss of hydroxyproline and calcium and by the extension of resorption lacunae. E-64, a specific inhibitor of cysteine proteinases, inhibits reversibly the resorption of cultured bones without affecting the hormone-induced secretion of lysosomal hydrolases. Given in vivo to rats, the proteinase inhibitors, E-64 and leupeptin, both induce a concomitant fall in the serum calcium level and in the urinary excretion of hydroxyproline. These results provide evidence that cysteine proteinases, possibly lysosomal cathepsins, are necessary for bone resorption.

Animals↗
PubMed1994

The gene encoding the kappa-carrageenase of Alteromonas carrageenovora is related to beta-1,3-1,4-glucanases.

The nucleotide (nt) and deduced amino acid (aa) sequences are reported for the structural gene cgkA encoding kappa-carrageenase (M(r) 44,412) from the marine bacterium Alteromonas carrageenovora (ATCC 43555), a hydrolase involved in the degradation of kappa-carrageenan. The cgkA gene codes for a protein of 397 aa, with a signal peptide of 25 aa. The enzyme is a new member of family 16 of glycosyl hydrolases, which comprises beta-1,3-1,4-glucanases from various sources and the beta-agarase of Streptomyces coelicolor. It is proposed that residue Glu163 in the kappa-carrageenase from A. carrageenovora and Glu155 in the beta-agarase from S. coelicolor are important for catalysis.

Amino Acid Sequence↗
PubMed1977

Glycosphingolipid hydrolases: properties and molecular genetics.

This is a review of the properties and molecular genetics of six lysosomal hydrolases: beta-galactosidase, hexosaminidases A and B, alpha-galactosidase, beta-glucosidase and alpha-fucosidase. Each enzyme is discussed with regards to isoenzymes and substrate specificity, subunit structure, genetic relationship of isoenzymes and genetic variants. The molecular genetics of human diseases caused by deficiencies of each enzyme are discussed.

Animals↗
PubMed2003

Cloning and sequence analysis of endoglucanase genes from an industrial fungus, Aspergillus kawachii.

Three endoglucanase genes (cel5A, cel5B, and cel61A) were cloned from an industrial fungus, Aspergillus kawachii. Yeasts transformed with these cDNAs showed endoglucanase activity in medium. Cel5A and Cel61A contained a type 1 cellulose-binding domain (CBD1) at the C-terminus of the enzyme. The putative catalytic regions of Cel5A and Cel5B showed homology with various endoglucanases belonging glycosyl hydrolase family 5 (GH5). Cel5B showed high homology with Cel5A in catalytic region, but it lacked CBD1 and linker. The cel5A contained four introns, whereas cel5B contained five introns. The putative catalytic region of Cel61A showed homology with enzymes belonging to GH61. The cel61A contained no introns.

Amino Acid Sequence↗
PubMed1996

Synthesis of a glioma-related ganglioside, O-Ac GM3 having 3-O-Ac ceramide and its substrate property toward hydrolases.

An O-acetyl group was selectively introduced into the ceramide moiety at the C-3-O on ganglioside GM3 containing N-acetyl neuraminic acid, the product of which has been previously found in rat glioma tissue as a glioma-associated ganglioside. The introduction of the acetyl residue involved a two-step process involving per O-acetylation of GM3 and saponification with a mild alkaline solution in a bilayer system constituted of water and water-immiscible organic solvent. Of the several solvents studied, 2-pentanol and diethyl ether gave the highest yields (68% and 62%, respectively). The chemical structure of the synthesized 3-O-acetyl GM3 was confirmed by proton nuclear magnetic resonance spectroscopy and fast atom bombardment-mass spectrometry, as well as by comparing the mobilities on thin-layer chromatography of its exoglycosidase-digested products with those of the synthesized, authentic 3-O-acetyl-lactosylceramide and ceramide. Furthermore, the substrate specificities of both 3-O-acetyl GM3 and 3-O-acetyl sphingomyelin toward exo- and endo-hydrolases were examined, revealing that they were hardly cleaved by the endoglycoceramidase and sphingolipid N-deacylase for the 3-O-acetyl GM3 and by sphingomyelinase for 3-O-acetyl sphingomyelin. Thus, the enzymes were found to recognize a free C-3 hydroxyl group on ceramide.

Acetylation↗
PubMed2002

Probing the catalytically essential residues of the alpha-L-arabinofuranosidase from Thermobacillus xylanilyticus.

The alpha-L-arabinofuranosidase D3 from Thermobacillus xylanilyticus is an arabinoxylan-debranching enzyme which belongs to family 51 of the glycosyl hydrolase classification. Previous studies have indicated that members of this family are retaining enzymes and may form part of the 4/7 superfamily of glycosyl hydrolases. To investigate the active site of alpha-L-arabinofuranosidase D3, we have used sequence alignment, site-directed mutagenesis and kinetic analyses. Likewise, we have shown that Glu(28), Glu(176) and Glu(298) are important for catalytic activity. Kinetic data obtained for the mutant Glu(176)-->Gln, combined with the results of chemical rescue using the mutant Glu(176)-->Ala, have shown that Glu(176) is the acid-base residue. Moreover, NMR analysis of the arabinosyl-azide adduct, which was produced by chemical rescue of the mutant Glu(176)-->Ala, indicated that alpha-L-arabinofuranosidase D3 hydrolyses glycosidic bonds with retention of the anomeric configuration. The results of similar chemical rescue studies using other mutant enzymes suggest that Glu(298) might be the catalytic nucleophile and that Glu(28) is a third member of a catalytic triad which may be responsible for modulating the ionization state of the acid-base and implicated in substrate fixation. Overall, these findings support the hypothesis that alpha-L-arabinofuranosidase D3 belongs to the 4/7 superfamily and provide the first experimental evidence concerning the catalytic apparatus of a family 51 arabinofuranosidase.

Amino Acid Sequence↗
PubMed1996

Regional distribution and hormonal control of estrogen-dependent oviduct-specific glycoprotein messenger ribonucleic acid in the baboon (Papio anubis).

Our objective in this study was to complete the sequence of the baboon oviductal glycoprotein, examine the hormonal regulation of the oviductal glycoprotein mRNA, and determine whether there was a regional variation within the oviduct in the level of oviductal glycoprotein mRNA expression. Finally, because of the structural similarity of the amino terminal end of the oviductal glycoprotein to chitinases, we sought to determine whether the oviductal glycoprotein functions as a glycosyl hydrolase. The total transcript length of the baboon oviductal glycoprotein was determined to be 2228 nucleotides in length plus a poly(A) tail. The largest open reading frame was 623 amino acids, which would produce a protein of 69.3 kDa. The first 420 amino acids were highly homologous to the amino acid sequence of other oviductal glycoproteins, but the remainder of the sequence differed considerably from that of all other species except the human. Although the N-terminal region exhibited sequence similarity to chitinases, the oviductal glycoprotein did not exhibit any activity towards typical chitinase substrates. The oviductal glycoprotein mRNA levels were elevated to approximately the same extent in the fimbria, ampulla, and isthmus of the oviduct after estradiol treatment and in the late follicular stage of the menstrual cycle. The oviductal glycoprotein mRNA levels were lower in the early follicular stage and early luteal stage and were not detectable in the late luteal stage or in progesterone-treated baboons. These results indicate that the oviductal glycoprotein mRNA is induced by estradiol and is present at the highest levels at the time of fertilization. Although there is structural homology with chitinases, no such glycosyl hydrolase activity could be detected. However, the common structure of the N-terminal region of the oviductal glycoproteins implies that it has the same, as yet unknown, function in all species.

Amino Acid Sequence↗
PubMed1976

Highly specific bacteriophage-associated polysaccharide hydrolases for Klebsiella aerogenes type 8.

Two phage-bound polysaccharide hydrolases specific for Klebsiella aerogenes type 8 exopolysaccharides were isolated. Each enzyme was specific for the polysaccharide produced by the host strain. One enzyme hydrolysed a pyruvylated and acetylated polymer, while the other only acted on the substrate lacking these substituents. Both enzymes were endogalactosidases releasing tetrasaccharides from their substrates which were only hydrolysed to a limited extent.

Acetates↗
PubMed1993

Ca2+ deficiency, selective alpha-glucosidehydrolase inhibition, and insulin secretion.

We investigated the relation between activities of islet glycogenolytic alpha-glucosidehydrolases and insulin secretion induced by glucose and 3-isobutyl-1-methylxanthine (IBMX) by means of suppressing 1) insulin release (Ca2+ deficiency) and 2) islet alpha-glucosidehydrolase activity (selective inhibition by the deoxynojirimycin derivative miglitol). Additionally, the in vivo insulin response to both secretagogues was examined. We observed that, similar to glucose-induced insulin release, islet glycogenolytic hydrolases (acid amyloglucosidase, acid alpha-glucosidase) were highly Ca2+ dependent. Acid phosphatase, N-acetyl-beta-D-glucosaminidase, or neutral alpha-glucosidase (endoplasmic reticulum) was not influenced by Ca2+ deficiency. In Ca2+ deficiency IBMX-induced insulin release was unaffected and was accompanied by reduced activities of islet alpha-glucosidehydrolases. Miglitol strongly inhibited glucose-induced insulin release concomitant with a marked suppression of islet alpha-glucosidehydrolase activities. Direct addition of miglitol to islet homogenates suppressed acid amyloglucosidase [half-maximal effective concentration (EC50) approximately 10(-6) M] and acid alpha-glucosidase. Acid phosphatase and N-acetyl-beta-D-glucosaminidase were unaffected. The miglitol-induced inhibition of glucose-stimulated insulin release was dose dependent (EC50 approximately 10(-6) M) and displayed a remarkable parallelism with the inhibition curve for acid amyloglucosidase. The in vivo insulin secretory response to glucose was markedly reduced in dystrophic mice (low amyloglucosidase), whereas the response to IBMX was unaffected. In summary, islet glycogenolytic hydrolases are Ca2+ dependent, and acid amyloglucosidase is directly involved in the multifactorial process of glucose-induced insulin release. In contrast the mechanisms of IBMX-stimulated insulin secretion operate independently of these enzymes. The effects of miglitol, a drug currently used in diabetes therapy, deserves further investigation.

1-Deoxynojirimycin↗
PubMed1975

Enzymes of the human intestinal brush border membrane. Identification after gel electrophoretic separation.

The postition of a number of human intestine brush border membrane enzyme activities in polyacrylamide gels after electrophoresis has been determined. These activities are, in order from the origin, maltase/glucoamylase, lactase/phlorizin hydrolase, maltase/sucrase/isomaltase, enteropeptidase, trehalase and gamma-glutamyl-transferase. Leucylnaphthylamide hydrolyzing activity was inactivated by sodium dodecylsulfate and its position was not determined. The positions of the activities have been correlated with the positions of protein bands previously determined. One such band situated between enteropeptidase and alkaline phosphatase has not been identified.

Alkaline Phosphatase↗
PubMed1997

Action patterns and mapping of the substrate-binding regions of endo-(1-->5)-alpha-L-arabinanases from Aspergillus niger and Aspergillus aculeatus.

The substrate binding sites of endo-(1-->5)-alpha-L-arabinanases (EC 3.2.1.99) from Aspergillus niger and Aspergillus aculeatus were investigated using reduced and regular (1-->5)-alpha-L-arabino-oligosaccharides and high performance anion exchange chromatographic analysis. Calculation of bond cleavage frequencies and kcat/K(m) parameters for these substrates enabled the determination of the number of arabinofuranosyl binding subsites and the estimation of the binding affinities of each subsite. The A. aculeatus endo-arabinanase has six subsites arranged symmetrically around the catalytic site, while the A. niger endo-arabinanase has five subsites; two from the catalytic site towards the non-reducing end of the bound substrate and three toward the reducing end. The two subsites directly adjacent to the catalytic sites in both the A. niger and A. aculeatus endo-arabinanase have near-zero net free energy of binding. These results are unlike most glycopyranosyl endo-hydrolases studied which have net negative (unfavourable) energies of interaction at these two subsites, and may be related to the greater conformational flexibility of arabinofuranosyl residues than glycopyranosyl residues. The complete subsite maps are also rationalized with regard to the observed action patterns of these enzymes on linear (1-->5)-alpha-L-arabinan.

Aspergillus↗
PubMed1987

Acrosomal enzymes and ultrastructure of unfrozen and cryotreated human spermatozoa.

Pooled semen judged to be normal in all parameters was divided into a number of aliquots which were either 1) kept untreated; 2) mixed with glycerol (10% v/v); 3) washed by centrifugation and resuspended to the original volume with buffer; or 4) washed and resuspended in buffer with glycerol (10% v/v). The progressive motility, viability, ultrastructure, and acrosomal enzyme activity (8 different hydrolases) were studied before and after cryotreatment. The described washing procedure effectively removed seminal plasma, and did not alter sperm motility, sperm viability, sperm ultrastructure, or the acrosomal enzymes studied. Glycerol (10%, v/v) had a deleterious effect on most parameters evaluated before cryotreatment. Cryotreatment severely altered the motility and viability of the spermatozoa and their acrosomal morphology but did not cause significant decreases in most of the acrosomal hydrolases measured. However, acrosin/proacrosin levels decreased by 50-80% and were correlated to the acrosomal damage. A simple assay for the measurement of acrosin/proacrosin enzyme levels in whole sperm is presented which could be used as a monitor for acrosomal integrity. No significant differences were seen between the samples cryotreated in the absence or presence of seminal plasma.

Acid Phosphatase↗