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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

High-sensitivity differential scanning calorimetry has been applied to characterize the irreversible thermal denaturation of a cellulase, assuming that thermal denaturation takes place according to the kinetic scheme N-k-->D, where k is a first-order kinetic constant that changes with temperature, as given by the Arrhenius equation; N the native state, and D the denatured one. On the basis of this model, the values of the rate constant as a function of temperature and the activation energy were calculated. The analytical data obtained with the fluorescence method as well by measurement of the enzymatic activity temperature dependence support this two-state kinetic model.
Biochem Mol Biol Int 1996 Feb
PMID:Differential scanning calorimetry of the irreversible thermal denaturation of cellulase from Streptomyces halstedii JM8. 893 30

Endoglucanase I (EG I) is a cellulase, from glycosyl hydrolase family 7, which cleaves the beta-1,4 linkages of cellulose with overall retention of configuration. The structure of the EG I from Fusarium oxysproum, complexed to a nonhydrolyzable thiooligosaccharide substrate analogue, has been determined by X-ray crystallography at a resolution of 2.7 A utilizing the 4-fold noncrystallographic symmetry present in the asymmetric unit. The electron density map clearly reveals the presence of three glucosyl units of the inhibitor, consistent with the known number of sugar-binding subsites, located at the active site of the enzyme in the -2, -1, and +1 subsites, i.e., actually spanning the point of enzymatic cleavage. The pyranose ring at the point of potential enzymatic cleavage is clearly distorted from the standard 4C1 chair as was originally suggested for beta-retaining enzymes by Phillips [Ford, L.O., Johnson, L.N., Machin, P. A., Phillips, D.C., & Tijan, T. (1974) J. Mol. Biol, 88, 349-371]. The distortion observed goes beyond the "sofa" conformation observed in previous studies and results in a conformation whose salient feature is the resulting quasi-axial orientation for the glycosidic bond and leaving group, as predicted by stereoelectronic theory. An almost identical conformation has recently been observed in a complex of chitobiase with its unhydrolyzed substrate [Tews, I., Perrakis, A., Oppenheim, A., Dauter, Z., Wilson, K. S., & Vorgias, C. E. (1996) Nat. Struct. Biol. 3, 638-648]. The striking similarity between these two complexes extends beyond the almost identical pyranose ring distortion. The overlap of the two respective sugars places the enzymatic nucleophile of endoglucanase I in coincidence with the C2 acetamido oxygen of N-acetylglucosamine in the catalytic site of the chitobiase, substantiating the involvement of this group in the catalytic mechanism of chitobiase and related chitinolytic enzymes. The endoglucanase I complex with the thiosaccharide substrate analogue clearly illustrates the potential of nonhydrolyzable sulfur-linked oligosaccharides in the elucidation of substrate binding and catalysis by glycosyl hydrolases.
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PMID:Structure of the Fusarium oxysporum endoglucanase I with a nonhydrolyzable substrate analogue: substrate distortion gives rise to the preferred axial orientation for the leaving group. 895 78

Functional analysis of the cellulase promoter cbh1 of the filamentous fungus Trichoderma reesei was carried out using the Escherichia coli lacZ gene as a reporter. An assay based on cultivation on solid medium in microtiter plates was developed that allows rapid and reliable semiquantitative analysis of beta-galactosidase expression of a large number of transformants. A series of deletions and specifically designed alterations were made covering 2.2 kb of the cbh1 promoter. Removal of sequences upstream of nucleotide -500 in relation to the initiator ATG abolished glucose repression. Mutation of a single hexanucleotide sequence 5'GTGGGG at nucleotide -720 was sufficient for derepression. This site is similar to the binding sites of the glucose repressors MIG1 of Saccharomyces cerevisiae and CREA/CREI of filamentous fungi. Removal of the glucose repressor site did not affect sophorose induction. Sophorose induction of the promoter was retained even in deletion derivatives lacking sequences upstream of position -161, which retained about 70 bp upstream of the transcription start point and only 30 bp upstream of the TATA box.
Mol Gen Genet 1996 Dec 13
PMID:Functional analysis of the cellobiohydrolase I promoter of the filamentous fungus Trichoderma reesei. 954 39

Cellulase and xylanase cDNAs were isolated from a cDNA library of the polycentric anaerobic fungus Orpinomyces sp. strain PC-2 constructed in Escherichia coli. The cellulase cDNA (celB) was 1.8 kb long with an open reading frame (ORF) coding for a polypeptide of 471 amino acids, and the xylanase cDNA (xynA) was 1.2 kb long with an ORF encoding a polypeptide of 362 amino acids. Single transcripts of 1.9 kb for celB and 1.5 kb for xynA were detected in total RNA of Orpinomyces grown on Avicel. Genomic DNA regions coding for CelA and XynA were devoid of introns. The enzymes were highly homologous (80 to 85% identity) to the corresponding enzymes of the monocentric anaerobic fungus Neocallimastix patriciarum and, like those, contained in addition to a catalytic domain, a noncatalytic repeated peptide domain (NCRPD). The Orpinomyces xylanase contained one catalytic domain and thus differed from the Neocallimastix xylanase, which had two similar catalytic domains (H. J. Gilbert, G. P. Hazlewood, J. I. Lauie, C. G. Orpin, and G. P. Xue, Mol. Microbiol. 6:2065-2072, 1992). Two peptides corresponding to the catalytic domain and the NCRPD of XynA were synthesized, and antibodies against them were raised and affinity column purified. The antibodies against the catalytic domain peptide reacted specifically with the xylanases of Orpinomyces and Neocallimastix, while the antibodies against the NCRPD reacted with many (at least eight) extracellular proteins of Orpinomyces and Neocallimastix, suggesting that the NCRPD is present in a number of polypeptides.
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PMID:Monocentric and polycentric anaerobic fungi produce structurally related cellulases and xylanases. 902 40

The endo-beta-1,4-glucanases, or cellulases, of higher plants are cell wall-associated enzymes believed to function in cell wall changes associated with the diverse processes of fruit ripening, organ abscission and cell elongation. We have isolated and characterized cDNA and genomic clones encoding a cellulase, PCEL1, which is abundant in ripening pepper fruit. Genomic analysis indicates that PCEL1 is encoded by a single gene, PCEL1, which belongs to a small, structurally divergent gene family. In ripening fruit, PCEL1 transcription is initiated at two distinct sites which yields overlapping mRNA species of 1.7 and 2.1 kb. High-level accumulation of both transcripts occurs in red fruit, while the 1.7 kb transcript is detected at a much lower level in stem and petiolar tissue. The increase in cellulase activity which is measured during fruit ripening is the product of PCEL1 expression and is tightly coupled to fruit reddening. High-level applications of ethylene serve to enhance the rate of ripening and the accumulation of PCEL1 mRNA. A direct role for ethylene in regulating PCEL1 expression is shown by the exclusive induction, in immature green fruit, of the 1.7 kb transcript in response to prolonged high-level exposure to ethylene--a pattern of expression not observed in fruit development on the vine.
Plant Mol Biol 1997 Jan
PMID:Isolation and characterization of a gene encoding endo-beta-1,4-glucanase from pepper (Capsicum annuum L.). 903 58

At the host-pathogen interface of hyphae penetrating host cell walls in the rye ovary, a lack of cellulase-gold labeling of beta-1, 4-glucan in host cell walls indicates that enzymatic degradation of cellulose might be an important factor during the infection of rye by Claviceps purpurea. Using cbh1 from Trichoderma reesei as a probe, a putative cellulase gene (cel1) was isolated from a genomic library of the C. purpurea strain T5. The coding region of 1,616 bp contains two introns and a putative signal peptidase cleavage site, leaving a coding capacity of 437 amino acids for the mature protein. The derived amino acid sequence shares significant homology with other fungal cellobiohydrolases and lacks the substrate binding domain. Expression analysis using reverse transcriptase-polymerase chain reaction (RT-PCR) shows that cel1 is induced during the first days of infection of rye by C. purpurea. It may be involved in the penetration and degradation of host cell walls by depolymerizing plant beta-1, 4-glucan and, therefore, play a role in the infection process.
Mol Plant Microbe Interact 1997 Mar
PMID:Cel1, probably encoding a cellobiohydrolase lacking the substrate binding domain, is expressed in the initial infection phase of Claviceps purpurea on Secale cereale. 905 32

A mutant of Erwinia carotovora subsp. carotovora, AH2552, created by a Mud1 insertion was found to be reduced in plant pathogenicity and deficient in extracellular protease and cellulase activity, although it produced normal levels of pectate lyase and polygalacturonase. A cosmid clone, pEC462, was isolated from a wild-type E. carotovora subsp. carotovora DNA library that concomitantly restored pathogenicity and protease and cellulase activities of AH2552 to wild-type levels when present in trans. The genetic locus that was disrupted in AH2552 by insertion of Mud1 has been designated rpfA, for regulator of pathogenicity factors. Sequencing of the rpfA region identified an open reading frame of 2,787 bp, and the predicted 929-amino acid polypeptide shared high identity with several two-component sensor-regulator proteins: BarA from Escherichia coli, ApdA from Pseudomonas fluorescens, PheN from P. tolaasii, RepA from P. viridiflava, LemA from P. syringae pv. syringae, and RpfC from Xanthomonas campestris pv. campestris. The RpfA locus described in this study encodes a putative sensor kinase protein that is involved in both extracellular protease and cellulase production and the pathogenicity of E. carotovora subsp. carotovora on potato tubers.
Mol Plant Microbe Interact 1997 Apr
PMID:Identification of a pathogenicity locus, rpfA, in Erwinia carotovora subsp. carotovora subsp. carotovora that encodes a two-component sensor-regulator protein. 910 Mar 85

The intact and the truncated endo-beta-1,4-glucanase expressed in Bacillus megaterium by a B. subtilis gene were purified and its adsorption characteristics to cellulosic materials were investigated. The intact enzyme was purified by affinity towards insoluble cellulose and Mono-Q chromatography. The truncated enzyme was purified by gel filtration and chromatofocusing. The molecular activities of these enzymes towards the soluble substrates were identical. Optimum pH and temperature of these two types of enzyme were same, 5.5 and 60 degrees C, respectively. However, the insoluble substrate, Avicel, was hydrolyzed slowly by the intact endoglucanase while the truncated endoglucanase did not hydrolyze the Avicel. Isoelectric points of the intact and the truncated enzyme were 6.2 and 5.6, respectively. In a Sephadex column the large form of intact endoglucanase was eluted later than the small form of truncated enzyme. Only the intact enzyme was strongly adsorbed to Avicel. The practical maximum adsorption was about 20 mg/g of Avicel at pH 6.0 and 4 degrees C.
Biochem Mol Biol Int 1997 Apr
PMID:Adsorption of Bacillus subtilis endo-beta-1,4-glucanase to cellulosic materials. 911 28

Two endo-beta-1,4-glucanase components (YEG1 and YEG2) of the endogenous cellulase from the Japanese subterranean termite, Reticulitermes speratus, were purified to homogeneity using gel filtration and hydroxylapatite chromatography, and their enzymatic properties were investigated. Using sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE), YEG1 and YEG2 had M(r) of 42 kDa and 41 kDa, respectively. Both components had an optimal pH of 6.0, an optimal temperature of 50 degrees C and were stable at 40 degrees C for at least 30 min. Both components showed high activity on sodium carboxymethylcellulose (CMC), 73.6 U/mg protein for YEG1 and 83.4 U/mg protein for YEG2. The K(m) values of YEG1 and YEG2 on CMC were 1.83 mg/ml and 1.48 mg/ml, respectively. YEG1 did not hydrolyse cellotetraose or cellotriose, whereas YEG2 hydrolysed cellotetraose to cellobiose and cellotriose to cellobiose and glucose. Both YEG1 and YEG2 hydrolysed cellopentaose to cellotriose and cellobiose. Neither component hydrolysed cellobiose. The hydrolytic products from crystalline cellulose (Sigmacell type 20) by YEG1 and YEG2 were cellobiose and a trace amount of glucose. Polyclonal mouse anti-serum raised against YEG2 crossreacted with YEG1, suggesting a common origin for both components. Using this anti-serum, Western blotting and immunohistochemistry showed the presence of YEG1 and YEG2 in the salivary glands, but not in the midgut epithelium. The data suggest that the salivary glands are the site of secretion of endo-beta-1,4-glucanase in R. speratus.
Insect Biochem Mol Biol 1997 Apr
PMID:Site of secretion and properties of endogenous endo-beta-1,4-glucanase components from Reticulitermes speratus (Kolbe), a Japanese subterranean termite. 913 11

The cellulase complex from Trichoderma reesei was polyethylene glycol (PEG)-modified with considerable retention of endo-beta-1,4-glucanase activity, as evaluated by the carboxymethylcellulase (CMCase) assay. While resistance towards heat denaturation was the same for either form, susceptibility towards proteolysis was slightly greater for the PEG-conjugate, in contrast to most reports using other enzymes. The circulatory life of endoglucanase activity associated with the complex was enhanced upon PEG-modification. This was confirmed by demonstrating degradation of chromogen-labelled substrate injected (i.v.) after 24hr of administration of the PEG-ylated enzymes; in contrast the substrate remained undegraded in mice pretreated with the native complex. The PEG-modified complex could be a good candidate for the treatment of pneumoconioses of textile workers exposed to cotton dust.
Biochem Mol Biol Int 1997 Jun
PMID:Enhanced in vivo catalytic activity of PEG-modified cellulase complex from Trichoderma reesei. 919 89


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