Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.25.1 (proteasome)
 28,817 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

During the course of the transformation of 1,3-dichloro-2-propanol (DCP) into (R)-3-chloro-1,2-propanediol [(R)-MCP] with the cell extract of Corynebacterium sp. strain N-1074, epichlorohydrin (ECH) was transiently formed. The cell extract was fractionated into two DCP-dechlorinating activities (fractions Ia and Ib) and two ECH-hydrolyzing activities (fractions IIa and IIb) by TSKgel DEAE-5PW column chromatography. Fractions Ia and Ib catalyzed the interconversion of DCP to ECH, and fractions IIa and IIb catalyzed the transformation of ECH into MCP. Fractions Ia and IIa showed only low enantioselectivity for each reaction, whereas fractions Ib and IIb exhibited considerable enantioselectivity, yielding R-rich ECH and MCP, respectively. Enzymes Ia and Ib were isolated from fractions Ia and Ib, respectively. Enzyme Ia had a molecular mass of about 108 kDa and consisted of four subunits identical in molecular mass (about 28 kDa). Enzyme Ib was a protein of 115 kDa, composed of two different polypeptides (about 35 and 32 kDa). The specific activity of enzyme Ib for DCP was about 30-fold higher than that of enzyme Ia. Both enzymes catalyzed the transformation of several halohydrins into the corresponding epoxides with liberation of halides and its reverse reaction. Their substrate specificities and immunological properties differed from each other. Enzyme Ia seemed to be halohydrin hydrogen-halide-lyase which was already purified from Escherichia coli carrying a gene from Corynebacterium sp. strain N-1074.
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PMID:Resolution and some properties of enzymes involved in enantioselective transformation of 1,3-dichloro-2-propanol to (R)-3-chloro-1,2-propanediol by Corynebacterium sp. strain N-1074. 144 32

A cDNA clone isolated from an Arabidopsis thaliana cell suspension culture library showed considerable similarities to the proteasome 28 kDa alpha subunit of Drosophila [(1990) Gene 90, 235-241]. The 250 amino acid-long protein encoded by Arabidopsis TAS-g64 clone has important homologies in its primary structure and in the predicted secondary structure with the PROS-28.1 clone from Drosophila. The only divergence observed between the two sequences is for the 20 C-terminal amino acids. This subunit might share important functions in both kingdoms, as revealed by the important conservation between plants and animals. In plant cells it is encoded by a single-copy gene and probably regulated by stress and/of division.
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PMID:Cloning and sequence analysis of a cDNA clone from Arabidopsis thaliana homologous to a proteasome alpha subunit from Drosophila. 151 3

Monoclonal antibodies (mAbs) were generated to proteasome purified from human erythrocytes. Five of six proteasome-specific mAbs reacted with three subunits in the molecular mass range of 25-28 kDa, indicating a common epitope. The other mAb (AP5C10) exhibited a more restricted reactivity, recognizing a 32-kDa subunit of the proteasome purified in its latent state. However, when the proteasome is isolated in its active state, AP5C10 reacts with a 28-kDa subunit, evidence for processing of the proteasome subunits during purification. Purified proteasome preparations which exhibited partial latency have both AP5C10 reactive subunits. Although the 32-kDa subunit appears required for latency, loss of this component and generation of the 28-kDa component are not obligatory for activation. The 32- and 28-kDa subunits can each be further resolved into three components by isoelectric focusing. The apparent loss of 4 kDa during the conversion of the 32- to 28-kDa subunit is accompanied by a shift to a more basic pI for each polypeptide. Western blots of the early steps of proteasome purification reveal an AP5C10-reactive protein at 41 kDa. This protein was separated from proteasomes by sizing chromatography and may represent a pool of precursor subunits. Since the 32-kDa subunit appears necessary for latency, it is speculated to play a regulatory role in ATP-dependent proteolytic activity.
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PMID:A monoclonal antibody that distinguishes latent and active forms of the proteasome (multicatalytic proteinase complex). 155 7

Six basic proteins of 26 to 38 kDa with isoelectric points (pI) > or = 8.5 were abundant in proteins separated by two-dimensional SDS-PAGE from adult rat peritoneal mast cells (MC). One was identified previously as rat mast cell proteinase (RMCP) 1, a chymase of 26 to 28 kDa, pI > 9.0. Microsequence analyses showed that two polypeptides of about 29 and 30 kDa had NH2 terminal amino acid sequences homologous to mouse MC proteinase 5 (MCP-5), whereas the amino terminals of the 33, 35, and 36 kDa proteins were homologous to MC carboxypeptidase A (MC-CPA). Rabbit Abs produced against synthetic peptides of the identified NH2 terminal sequences were used in immunoblot studies. At least three proteins reacted with Abs to MC-CPA, whereas Abs to MCP-5 detected three adjacent polypeptides, rather than just the two identified by using microsequence analysis. Removal of oligosaccharide side chains using peptide:N-glycosidase F reduced the heterogeneity of each set of three polypeptides (MCP-5 and MC-CPA) to a band of each protein of a lower M(r). The serine proteinase inhibitor [3H]diisopropylfluorophosphate ([3H]DFP) bound to a proteinase of 30 to 35 kDa, which is probably MC tryptase (pI < or = 6.0). Immunoblot analysis of proteins from intestinal mucosal mast cells showed RMCP-2, but not RMCP-1, MCP-5, or MC-CPA. This is the first report of MCP-5 in the rat and of clearly distinguishable glycosylated forms of MC CPA. These proteinases appear to be restricted in their distribution to selected MC populations, but little is known about their functions.
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PMID:Proteinases of rat mast cells. Peritoneal but not intestinal mucosal mast cells express mast cell proteinase 5 and carboxypeptidase A. 759 1

A previously undescribed high molecular mass protein (HMP) from human erythrocyte membranes was solubilized by Triton X-100 and purified on a calmodulin-agarose column in the presence of Ca2+. It was shown to have a native molecular mass of 522-560 kDa, comprised of a single subunit of a molecular mass of 28 kDa (p28). The protein is associated with the lipid bilayer rather than with the cytoskeletal component of the membrane. The purified HMP showed peptidase-hydrolyzing activity toward substrates containing hydrophobic amino acids at the P1 position of the P2-P1 cleavage site. The activity was inhibited by serine proteinase inhibitors (leupeptin, phenylmethansulfonyl fluoride) and chymotrypsin inhibitors in particular (chymostatin, N-tosyl-L-phenylalanine chloromethyl ketone). The enzyme exhibited maximal activity at slightly alkaline pH (7.5-8.5) and at 37 degrees C and was stimulated over a narrow range of SDS concentrations (maximal at 0.05%). HMP was found to cross-react in Western blots with an antibody raised against the rabbit multicatalytic proteinase. The single subunit of HMP therefore contains both the catalytic activity and a sequence necessary for its association into a multimeric complex. The properties of the human erythrocyte membrane HMP described indicate that it is a novel peptidase related to the ubiquitous multicatalytic proteinase.
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PMID:Characterization of a novel high molecular mass protein with peptidase activity purified from the human erythrocyte membrane by calmodulin affinity chromatography. 814 98

The mouse cDNA homologues of the rat C2, C9, and C5 subunits of the multicatalytic proteinase have been cloned and expressed in bacteria. The respective recombinant proteins were purified and used to produce specific anti-subunit antibodies. Immunoblotting of two-dimensional gels of purified rat liver multicatalytic proteinase showed that the C2 (32-kDa) and C9 (29-kDa) polypeptides are resolved into three and two isoelectric variants, respectively, likely due to post-translational modifications, i.e. phosphorylation, and the presence of two anti-C5 reacting polypeptides (25.5 and 23 kDa). Epitope mapping of the anti-C2-specific antibody with different constructs of the recombinant C2 protein allowed us to determine that one major epitope of this anti-C2 antibody is located within the last 9-11 amino acids of the C2 polypeptide. Affinity purified antibodies directed against the C2 COOH-terminal were able to discriminate the active and latent forms of the multicatalytic proteinase, supporting the conclusion that the C2 protein found in the active form of the enzyme is a polypeptide of 28 kDa, produced by the loss, at least, of the last 9-13 amino acids (DEPAEKADEPMEH) of the intact C2 (32-kDa) component. By in vitro treatment of the latent form of the enzyme with elastase, we show the conversion of the C2 (32-kDa) component to a 28-kDa protein with loss of recognition by the anti-C2 COOH-terminal affinity purified antibodies, but this limited degradation of the C2 component did not have any significant effect on the proteolytic activity (assayed with myelin basic protein and fluorogenic peptides) of the multicatalytic proteinase. It is suggested that the proteolytic cleavage of the C2 COOH-terminal region may be involved in the regulation of the interaction of the multicatalytic proteinase with other cellular proteins and/or in the translocation of the complex to the nucleus.
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PMID:Antibodies against the C2 COOH-terminal region discriminate the active and latent forms of the multicatalytic proteinase complex. 817 1

Cloned cDNA of genomic segment 10 of Bombyx mori cytoplasmic polyhedrosis virus (CPV) was placed downstream from the lambda PL promoter in expression plasmid pRC23 and expressed in Escherichia coli cells. A polypeptide of the same molecular weight (28 kDa) as natural polyhedrin was synthesized at the level of approximately 10% of total host cell protein. This polypeptide was identified as CPV polyhedrin (r-polyhedrin) after comparative studies. The r-polyhedrin did not form any crystalline structure in E. coli cells but instead accumulated in the form of an insoluble inclusion body, even though natural polyhedrin is known to form a crystalline matrix (polyhedra) in infected insect cells. The purified r-polyhedrin complex, like natural polyhedra, was not soluble in neutral or acidic buffer but soluble in alkaline buffer. Upon solubilization, the r-polyhedrin complex did not undergo proteolytic degradation, while natural polyhedra were digested into small peptides by the associated protease. Incubation of r-polyhedrin with natural polyhedra in alkaline buffer, however, degraded the r-polyhedrin, resulting in an identical profile of peptide products to that of natural polyhedra. These results indicate that even though r-polyhedrin molecules produced in E. coli cells are not in the natural conformation, the molecules can present the identical cleavage sites to the polyhedra-associated alkaline protease. Experiments showed that the alkaline protease was associated with the matrix of polyhedra and not with virus particles.
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PMID:Expression in Escherichia coli of the cloned polyhedrin gene of Bombyx mori cytoplasmic polyhedrosis virus. 828 55

We have identified two distinct subunits of 20 S proteasomes that are associated with RNase activity. Proteasome subunits zeta and iota, eluted from two-dimensional Western blots, hydrolysed tobacco mosaic virus RNA, whereas none of the other subunits degraded this substrate under the same conditions. Additionally, proteasomes were dissociated by 6 M urea, and subunit zeta, containing the highest RNase activity, was isolated by anion-exchange chromatography and gel filtration. Purified subunit zeta migrated as a single spot on two-dimensional PAGE with a molecular mass of approx. 28 kDa. Addition of anti-(subunit zeta) antibodies led to the co-precipitation of this proteasome subunit and nuclease activity. This is the first evidence that proteasomal alpha-type subunits are associated with an enzymic activity, and our results provide further evidence that proteasomes may be involved in cellular RNA metabolism.
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PMID:Involvement of proteasomal subunits zeta and iota in RNA degradation. 933 55

Activity of the 20S proteasome, which performs much of the cytosolic and nuclear proteolysis in eukaryotic cells, is controlled by regulatory complexes that bind to one or both ends of the cylindrical proteasome. One of these complexes, the 11S regulator (REG), is a complex of 28 kDa subunits that is thought to activate proteasomes toward the production of antigenic peptides. REG, purified from red blood cells, is a complex of REG alpha and REG beta subunits. We have crystallized recombinant REG alpha (rREG alpha) and collected diffraction data to 3.0 A resolution. The self-rotation function indicates that rREG alpha forms a heptameric ring in the crystal. Equilibrium sedimentation demonstrates that rREG alpha is a heptamer in solution also.
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PMID:The proteasome 11S regulator subunit REG alpha (PA28 alpha) is a heptamer. 938 52

A Streptomyces sp., which produces an alkaline protease inhibitor (API) exhibiting antifungal activity has been isolated from soil. The protein has been purified to homogeneity. The molecular characterization has revealed that it is a dimer (M(r) 28 kDa) with five disulphide linkages and has a pI of 3.8. API is a competitive type of inhibitor with a K(i) value of 2.5 x 10(-9) M. The inhibitor is stable over a pH range of 6 to 12 and a temperature range of 40 to 95 degrees C. API exhibits antifungal activity (in vitro) against phytopathogenic fungi such as Fusarium, Alternaria, and Rhizoctonia and also against Trichoderma, a saprophytic fungus. The antifungal activity of API appears to be associated with its ability to inhibit the fungal serine alkaline protease(s), which is indispensable for its growth. Retardation of the rate of fungal spore germination, as well as hyphal extention, was observed in the presence of API. Both the protease inhibitory and the antifungal activity were abolished on treatment of API with DTT (5 mM), suggestive of a common site for both the activities. This is the first report on API as a potential biocontrol agent against phytopathogenic fungi.
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PMID:Alkaline protease inhibitor: a novel class of antifungal proteins against phytopathogenic fungi. 1047 89


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