Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We present the three-dimensional structure of Trichoderma reesei endoglucanase 3 (Cel12A), a small, 218 amino acid residue (24.5 kDa), neutral pI, glycoside hydrolase family 12 cellulase that lacks a cellulose-binding module. The structure has been determined using X-ray crystallography and refined to 1.9 A resolution. The asymmetric unit consists of six non-crystallographic symmetry-related molecules that were exploited to improve initial multiple isomorphous replacement phasing, and subsequent structure refinement. The enzyme contains one disulfide bridge and is glycosylated at Asp164 by a single N-acetyl glucosamine residue. The protein has the expected fold for a glycoside hydrolase clan-C family 12 enzyme. It contains two beta-sheets, of six and nine strands, packed on top of one another, and one alpha-helix. The concave surface of the nine-stranded beta-sheet forms a large substrate-binding groove in which the active-site residues are located. In the active site, we find a carboxylic acid trio, similar to that of glycoside hydrolase families 7 and 16. The strictly conserved Asp99 hydrogen bonds to the nucleophile, the invariant Glu116. The binding crevice is lined with both aromatic and polar amino acid side-chains which may play a role in substrate binding. The structure of the fungal family 12 enzyme presented here allows a complete structural characterization of the glycoside hydrolase-C clan.
J Mol Biol 2001 Apr 27
PMID:The X-ray crystal structure of the Trichoderma reesei family 12 endoglucanase 3, Cel12A, at 1.9 A resolution. 1132 68

The characterization of a non-photosynthetic isoform of NADP-malic enzyme (NADP-ME) from maize roots, which represents nearly 7% of the total soluble protein of this tissue, was performed. The molecular properties of the purified protein, as well as the kinetic parameters determined, indicate that the NADP-ME isoform present in maize roots differs from the photosynthetic enzyme implicated in the C4 cycle, but is similar, or identical, to the enzyme previously characterized from etiolated maize leaves (Maurino, Drincovich and Andreo, Biochem. Mol. Biol. Int. 38 (1996) 239-250). A full-length ORF encoding a plastidic NADP-ME (almost identical to the maize root NADP-ME, GenBank accession number U39958) was cloned from a root cDNA library as well as isolated by reverse transcription (RT)-PCR using green leaves mRNA as template. These results indicate that root NADP-ME does not constitute a root-specific isoform, but represents a protein with a constitutive pattern of expression in plastids of the C4 plant maize. The amount of NADP-ME measured by activity, western and northern blot was modified when different stress conditions (including treatments with cellulase, fungal elicitors, jasmonate and hypoxic treatment) were applied to maize roots, indicating that the enzyme from maize roots is under transcriptional or post-transcriptional regulation by effectors related to plant defence responses. It is deduced that the induction of housekeeping genes, like non-photosynthetic NADP-ME, whose constitutive role may be the provision of reductive power in non-photosynthetic plastids, is likely to accompany the defence response.
Plant Mol Biol 2001 Mar
PMID:Non-photosynthetic 'malic enzyme' from maize: a constituvely expressed enzyme that responds to plant defence inducers. 1135 60

The alkaloid-rich latex of the opium poppy, Papaver somniferum L., is valued as a source of pharmaceuticals including thebaine, codeine, and morphine, but is also harvested for heroin production. The poppy laticifer system develops through the gradual disappearance of the common walls between differentiating laticifer elements throughout the plant. Gene homologues for cell-wall-degrading enzymes were found during random sequencing of an opium poppy latex cDNA library. RNA gel blot analysis of cellulase, polygalacturonase beta-subunit, 1,3-beta-glucanase, and xyloglucan endotransglycosylase homologues showed their expression was not limited to laticifers. In contrast, poppy gene homologues to pectin methylesterase (PME), pectin acetylesterase (PAE) and pectate lyase (PL) where all highly expressed and latex-specific. Enzyme assays confirmed the presence of PME, PAE, and PL activities in latex serum. The abundance of transcripts encoding pectin-degrading enzymes in latex suggests that these enzymes may play an important role in laticifer development.
Plant Mol Biol 2001 Mar
PMID:Expression and activity of cell-wall-degrading enzymes in the latex of opium poppy, Papaver somniferum L. 1141 15

The Arabidopsis KOR gene encodes a membrane-anchored endo-1,4-beta-D-glucanase involved in cell wall assembly. To obtain a more detailed knowledge of the small gene family encoding membrane-anchored endo-1,4-beta-D-glucanases in Arabidopsis thaliana, we have characterized two additional membrane-anchored endo-1,4-beta-D-glucanase genes. Sequence comparison indicates that KOR2 is distantly related to KOR and other plant membrane-anchored endo-1,4-beta-D-glucanases. The expression of KOR2 and KOR3 was followed by the beta-glucuronidase (gusA) reporter-gene method. While the KOR gene is most often expressed throughout the plant, KOR2::gusA and KOR3::gusA are active only in restricted cell types. We demonstrate that KOR2::gusA is expressed very early in the development of root hairs within the root differentiation zone (specialization zone) but not in the root-hair-bearing epidermal cells at the root/shoot junction (transition zone). Furthermore, KOR2::gusA is expressed in the proximal parts of leaves and floral organs (rosette and cauline leaves, sepals, petals and stamens), and in trichomes, as they develop at the tip of young leaves and later in more basal regions of the leaf blade. The KOR3::gusA construct is expressed in the trichome support cells that form a ring at the base of each trichome and in the bundle sheath cells which surround the vascular bundle within the leaf mesophyll tissue. Reverse transcription-polymerase chain reaction of Arabidopsis RNA confirmed the expression of KOR2::gusA and KOR3::gusA. In conclusion, although KOR2 and KOR3 have more restricted expression patterns than the previously characterized KOR gene, they are expressed in cell types at time points where cell wall assembly is likely to occur and, interestingly, differentially expressed in leaf trichomes and their support cells.
Plant Mol Biol 2001 Jun
PMID:Two Arabidopsis thaliana genes, KOR2 and KOR3, which encode membrane-anchored endo-1,4-beta-D-glucanases, are differentially expressed in developing leaf trichomes and their support cells. 1148 74

Erwinia chrysanthemi, a Gram-negative plant pathogen, secretes the cellulase Cel5 (formerly EGZ) via the type II secretion pathway (referred to as Out). Cel5 is composed of two domains, a large N-terminal catalytic domain (390 amino acid residues) and a small C-terminal cellulose-binding domain (62 amino acid residues) separated by a linker region. A combination of mutagenesis and structural analysis permitted us to investigate the structure/secretion relationships with respect to the catalytic domain of Cel5. The 3D structure of the catalytic domain was solved by molecular replacement at 2.3 A resolution. Cel5 exhibits the (beta/alpha)8 structural fold and two extra-barrel features. Our previous genetic study based upon tRNA-mediated suppression allowed us to predict positions of importance in the molecule in relation to structure and catalysis. Remarkably, all of the predictions proved to be correct when compared with the present structural information. Mutations of Arg57, which is located at the heart of the catalytic domain, allowed us to test the consequences of structural modifications on the secretion efficiency. The results revealed that secretability imposes remarkably strong constraints upon folding. In particular, an Arg-to-His mutation yielded a species that folded to a stable conformation close to, but distinct from the wild-type, which however was not secretable. We discuss the relationships between folding of a protein in the periplasm, en route to the cell exterior, and presentation of secretion information. We propose that different solutions have been selected for type II secreted exoproteins in order to meet the constraints imposed by their interaction with their respective secretion machineries. We propose that evolutionary pressure has led to the adaptation of different secretion motifs for different type II exoproteins.
J Mol Biol 2001 Jul 27
PMID:Type II protein secretion in gram-negative pathogenic bacteria: the study of the structure/secretion relationships of the cellulase Cel5 (formerly EGZ) from Erwinia chrysanthemi. 1150 95

The crystal structure of the catalytic domain of alkaline cellulase K was determined at 1.9 A resolution. Because of the most alkaliphilic nature and it's highest activity at pH 9.5, it is used commercially in laundry detergents. An analysis of the structural bases of the alkaliphilic character of the enzyme suggested a mechanism similar to that previously proposed for alkaline proteases, that is, an increase in the number of Arg, His, and Gln residues, and a decrease in Asp and Lys residues. Some ion pairs were formed by the gained Arg residues, which is similar to what has been found in the alkaline proteases. Lys-Asp ion pairs are disfavored and partly replaced with Arg-Asp ion pairs. The alkaline adaptation appeared to be a remodeling of ion pairs so that the charge balance is kept in the high pH range.
J Mol Biol 2001 Jul 27
PMID:Crystal structure of alkaline cellulase K: insight into the alkaline adaptation of an industrial enzyme. 1150 97

Previous dogma has maintained that cellulose, ingested by xylophagous or herbivorous animals, is digested by cellulolytic symbiotes. The first evidence in conflict with this contention involved the demonstration of cellulolytic activities in symbiote-free secreting organs (e.g., the salivary glands of termites) or defaunated guts. Following these demonstrations, possible endogenous cellulase components were purified from several cellulose-digesting invertebrates, but this research did little to change the general view concerning animal cellulose digestion. Thanks to recent developments in molecular biology, the existence of cellulases of animal origin has been firmly established. To date, cellulase genes have been reported from arthropods (insects and a crayfish) and nematodes. This paper describes and discusses the presence and nature of endogenous cellulases in higher animals.
Cell Mol Life Sci 2001 Aug
PMID:Animal cellulases. 1157 76

Cellobiohydrolase 58 (Cel7D) is the major cellulase produced by the white-rot fungus Phanerochaete chrysosporium, constituting approximately 10 % of the total secreted protein in liquid culture on cellulose. The enzyme is classified into family 7 of the glycosyl hydrolases, together with cellobiohydrolase I (Cel7A) and endoglucanase I (Cel7B) from Trichoderma reesei. Like those enzymes, it catalyses cellulose hydrolysis with net retention of the anomeric carbon configuration. The structure of the catalytic module (431 residues) of Cel7D was determined at 3.0 A resolution using the structure of Cel7A from T. reesei as a search model in molecular replacement, and ultimately refined at 1.32 A resolution. The core structure is a beta-sandwich composed of two large and mainly antiparallel beta-sheets packed onto each other. A long cellulose-binding groove is formed by loops on one face of the sandwich. The catalytic residues are conserved and the mechanism is expected to be the same as for other family members. The Phanerochaete Cel7D binding site is more open than that of the T. reesei cellobiohydrolase, as a result of deletions and other changes in the loop regions, which may explain observed differences in catalytic properties. The binding site is not, however, as open as the groove of the corresponding endoglucanase. A tyrosine residue at the entrance of the tunnel may be part of an additional subsite not present in the T. reesei cellobiohydrolase. The Cel7D structure was used to model the products of the five other family 7 genes found in P. chrysosporium. The results suggest that at least two of these will have differences in specificity and possibly catalytic mechanism, thus offering some explanation for the presence of Cel7 isozymes in this species, which are differentially expressed in response to various growth conditions.
J Mol Biol 2001 Dec 14
PMID:Family 7 cellobiohydrolases from Phanerochaete chrysosporium: crystal structure of the catalytic module of Cel7D (CBH58) at 1.32 A resolution and homology models of the isozymes. 1174 26

The phytopathogenic Pectobacterium chrysanthemi (Pch) PY35 secretes multiple isozymes of plant cell wall degrading enzyme cellulases. We cloned a second cel gene that encodes cellulase in Pch PY35. The inserted 2 kb fragment was subcloned in order to geneate pPY710 (cel8Y). The structural organization of the cel8Y gene consists of an open reading frame (ORF) of 999 bp that encodes 332 amino acid residues with a signal peptide of 23 amino acids. The predicted amino acid sequence of Cel8Y was very similar to that of Cellulomonas uda, but completely different from that of the Cel5Z of Pch PY35. It belonged to the glycoside hydrolase family 8, based on amino acid sequence similarities in contrast to Cel5Z of Pch PY35, which was confirmed as family 5. Cel8Y was not closely related to the known cellulases of Pectobacterium. It had the conserved region of the glycoside hydrolase family 8, ASDGDVLIAWALLKAGNKW. The apparent molecular mass of the Cel8Y protein was calculated to be approximately 34 kDa by a carboxymethylcellulosesodium dodecyl sulfate-polyacrylamide gel electrophoresis (CMC-SDS-PAGE). The Cel8Y had a calculated pl of 6.49. It was optimally active at pH 7 with an approximate optimal temperature around 40 degrees C. The cellulase activity of Cel8Y was lower than that of Cel5Z.
Mol Cells 2002 Feb 28
PMID:Cloning of the cel8Y gene from Pectobacterium chrysanthemi PY35 and its comparison to cel genes of soft-rot Pectobacterium. 1191 71

Lactose is at present the only soluble carbon source which can be used economically for the production by Hypocrea jecorina (= Trichoderma reesei) of cellulases or heterologous proteins under the control of cellulase expression signals. However, the mechanism by which lactose triggers the formation of cellulases is unknown. To enhance our understanding of lactose metabolism and its relationship to cellulase formation, we have cloned and characterized the gal7 gene (for galactose-1-phosphate uridylyltransferase) of H. jecorina. The gene encodes a polypeptide of 43.8 kDa, the sequence of which exhibits a moderate level of identity (about 50%) to that of the Gal7 proteins of Saccharomyces cerevisiae and Kluyveromyces lactis, and contains an active-site signature typical for galactose-1-phosphate uridylyltransferase family 1. H. jecorina gal7 is not clustered with other genes of galactose metabolism. A single 1.7-kb transcript is synthesized constitutively during the rapid growth phase and accumulated to twice this level during incubation in the presence of D-galactose and L-arabinose and the corresponding polyols (dulcitol, arabitol). A gal7 deletion mutant, constructed by replacing the gal7 reading frame by the H. jecorina pyr4 gene, was unable to grow on D-galactose between pH 4.5 and 7.5, thus proving that in H. jecorina gal7 is essential for metabolism of D-galactose, whereas the growth rate of the mutant on lactose was only reduced by about 50%. The rate of formation of cellobiohydrolase Cel7A and the abundance of the corresponding (cbh1) transcript during growth on lactose was only slightly lower in the absence of gal7, but a significant delay in decay of the cbh1 transcript was noted during later stages of growth. The results suggest that H. jecorina uses only the Leloir pathway for metabolism of D-galactose and lactose. Furthermore, we conclude that metabolism of lactose past the galactose-1-phosphate step is not essential for cellulase formation.
Mol Genet Genomics 2002 Mar
PMID:Lactose metabolism and cellulase production in Hypocrea jecorina: the gal7 gene, encoding galactose-1-phosphate uridylyltransferase, is essential for growth on galactose but not for cellulase induction. 1191 23


<< Previous 1 2 3 4 5 6 7 8 9 10