Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.24.11 (CD10)
9,792 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A new subunit, named RC6-I, of the rat 20 S proteasome was purified and the partial amino acid sequences of several peptide fragments obtained by digestion with lysyl-endopeptidase were determined by Edman degradation. Amplification of cDNAs encoding RC6-I by the polymerase chain reaction (PCR) technique revealed two types of cDNA, tentatively designated as RC6-IL and RC6-IS in order of size. The nucleotide sequences of the two cDNAs are identical except that RC6-IL contains an insertion of 18 nucleotides in the coding region compared with RC6-IS. The polypeptide predicted from the open reading frame of RC6-IS cDNA consists of 248 amino acid residues with a calculated molecular weight of 27,783. These values are consistent with those obtained by protein chemical analyses. Computer-assisted homology analysis showed that RC6-I belongs to the alpha-type subfamily of the proteasome gene family, which shows similarity to the alpha-subunit of the archaebacterium Thermoplasma acidophilum proteasome, and that the 18 nucleotide insert, encoding six amino acid residues, VVASVS, appears to be unique to RC6-IL, because this motif has not been conserved in any other alpha-type subunit. By reverse transcription (RT)-PCR analysis, the mRNAs for both RC6-IL and RC6-IS were found in all the rat tissues examined. These results suggest that proteasomes are present as a heterogeneous population, possibly for acquisition of diversity of functions.
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PMID:Molecular cloning of two types of cDNA encoding subunit RC6-I of rat proteasomes. 757 56

Proteinase B, an asparagine-specific endopeptidase, has been purified from germinating vetch (Vicia sativa L.) seeds. The final preparation consists of two enzymically active proteins with molecular masses of approximately 39 kDa and 37 kDa. Synthetic substrates were used to confirm cleavage specificity of the proteinase B preparation. As expected, the enzyme cleaves the substrates at the C-terminal side of Asn residues. The octapeptide ETRNGVEE was digested most efficiently. When Gly was replaced by Ile or Glu, cleavage took place with lower efficiency. Polyclonal antibodies displayed both proteins in cotyledon extracts of germinated vetch seeds. In addition, a strong cross-reacting protein band was found in cotyledon extracts of developing seeds, indicating the presence of a very similar enzyme during seed development. cDNA clones encoding proteinase B precursor have been obtained on the basis of the N-terminal amino acid sequence DDDFEGTRWAILLAGS, by means of the polymerase chain reaction. The cDNA clones contain an open reading frame of 1479 bp encoding a polypeptide of 493 amino acids. The precursor displayed 59% sequence identity to the cDNA-derived amino acid sequence of a vacuolar Asn-specific enzyme from the developing castor beam endosperm which is thought to catalyze the post-translational processing of pro-proteins into the mature forms. Proteinase B is synthesized de novo during seed germination. The results of Southern-blot analyses suggested that there are at least two genes for proteinase B.
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PMID:Purification, cDNA cloning and characterization of proteinase B, an asparagine-specific endopeptidase from germinating vetch (Vicia sativa L.) seeds. 770 62

Sixty-four kinds of cell lines were examined as to their ability to degrade glucagon using conditioned-media obtained from their protein-free cultures. Two human tumor cell lines were shown to produce this activity, and the cell line, HPC-YO, established from a human pancreatic carcinoma was shown to produce the highest level of activity. The glucagon-degrading enzyme (GDE) was purified from HPC-YO conditioned-medium by a combination of ion-exchange, gel filtration, and hydroxylapatite column chromatographies. The purified GDE also degraded vasoactive intestinal polypeptide (VIP) and secretin, however, it did not cleave EGF, gastrin, insulin, somatostatin, substance P, neurotensin, or growth hormone. The molecular weight of GDE is 83,000, as determined on SDS-polyacrylamide gel electrophoresis. The N-terminal amino acid sequence of GDE was blocked, and the five partial amino acid sequences obtained on lysyl-endopeptidase digestion were determined to be N-L-T-E-E-Y-D-V-S-D-G-E-I-E-L-L-Y-E-K, V-E-T-Y-Y-D-L-L-F-E-K, L-Y-W-F-L-D-E-A-K, S-N-S-T-S-Y-V-K, and Y-Y-A-S-T-S-Y-D-D-T-Y-K. The same or homologous amino acid sequences have not been found in known proteins, demonstrating that GDE is a novel peptidase that degrades the secretin family: glucagon, VIP, and secretin.
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PMID:A novel proteinase, glucagon-degrading enzyme, secreted by a human pancreatic cancer cell line, HPC-YO. 777 1

The acidic glucagon-degrading activity of hepatic endosomes has been attributed to membrane-bound forms of cathepsins B and D. Endosomal lysates processed full-length nonradiolabeled glucagon to 32 different peptides that were identified by amino acid analysis and full-length sequencing. These indicated C-terminal carboxypeptidase, endopeptidase as well as N-terminal tripeptidyl-aminopeptidase activities in endosomes. Glucagon proteolysis was inhibited 95% by E-64 and pepstatin A, inhibitors of cathepsins B and D, respectively. This was confirmed by the pH 6-dependent chemical cross-linking of [125I]iodoglucagon to a polypeptide of 30 kDa, which was immunodepleted by polyclonal anti-cathepsin B antibody, and the removal of greater than 80% of glucagon-degrading activity by polyclonal antibodies to cathepsins B and D. By similar criteria, insulin-degrading enzyme was ruled out as a candidate enzyme for endosomal proteolysis of glucagon. Lysosomal contamination was unlikely since all forms of cathepsin B in endosomes, i.e. the major 45-kDa inactive precursor as well as the lesser amounts of the 32- and 28-kDa active forms, were tightly bound to endosomal membranes. Furthermore the mature 29-kDa single-chain and 22-kDa heavy-chain forms of cathepsin L were undetectable in endosomes, although high levels of the 37-kDa proform were observed. Membrane association of the cathepsins B and D was not to the mannose 6-phosphate receptor since association was unaffected by mannose 6-phosphate and/or EDTA, thereby indicating a distinct endosomal receptor. Hence, a pool of active cathepsins B and D as well as a poorly defined tripeptidyl aminopeptidase is maintained in endosomes by selective membrane retention. These hydrolases degrade glucagon internalized into liver parenchyma early in endocytosis.
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PMID:Proteolysis of glucagon within hepatic endosomes by membrane-associated cathepsins B and D. 779 82

The cDNA libraries constructed from the human acute lymphoblastic leukemia cell line KM3 in the expression vector lambda gt11, were screened with the anti-CALLA (common acute lymphoblastic leukemia antigen) mAb (monoclonal antibody) J5. The selected J5-positive clone I containing a partial cDNA insert was isolated and sequenced. For completing the cDNA sequence the cDNA libraries were further screened by hybridization with the DIG (digoxigenin)-labelled DNA probe derived from clone I, the 5'-end region was analysed by 5'-RACE (rapid amplification of cDNA ends) using a sequence specific primer. In total a 1639 bp cDNA sequence was determined. The cDNA sequence contains a 1260 bp open reading frame and the untranslated 3'- and 5'-end sides. The 420 residue amino acid sequence, deduced from the cDNA sequence, unexpectedly differs fundamentally from CALLA (CD10) although clones I and II were J5-positive in immuno screening. The mature protein corresponding to the cDNA was isolated and characterized from the KM3 cells using polyclonal antisera raised against the in vitro expressed polypeptide from clone I. The protein is expressed on plasma membrane, in cytosol and is secreted into culture medium, its relative molecular mass was determined to be 55 kDa on SDS-PAGE. The deduced amino acid sequence from cDNA was confirmed by peptide sequences. The new protein contains a basic amino acid rich putative DNA binding domain (b) with a potential nuclear targeting signal, two helix-loop-helix (HLH) motif regions, concurrently EF-hand motifs, an acidic amino acid rich region (a) between the EF-hands, and a leucine zipper (Z) motif. This DNA binding protein therefore is characterized by a linked motif "b/HLH/a/HLH/Z". The protein was designated NEFA: DNA binding/EF-hand/acidic amino acid rich region.
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PMID:Human protein NEFA, a novel DNA binding/EF-hand/leucine zipper protein. Molecular cloning and sequence analysis of the cDNA, isolation and characterization of the protein. 781 91

Rabbit neutral endopeptidase 24.11 (NEP) is a type II membrane protein with a positively charged 27 amino acid residue NH2-terminal cytoplasmic domain, a 20 amino acid residue hydrophobic signal peptide/membrane anchor domain, and a large catalytic COOH-terminal domain exposed on the exoplasmic side of the membrane. To study the role of the cytosolic domain in anchoring NEP in the plasma membrane, we constructed two mutants in which this cytosolic domain was deleted. In the first mutant (NEP delta cyto), a Glu residue was present in NH2-terminus, while a Lys residue was substituted at the same position in the second mutant (NEP delta cyto(K)). To better understand the interaction of these mutants with the rough endoplasmic reticulum membrane, the mutated NEP cDNAs were transcribed and translated in vitro in the presence of microsomal membranes. Our studies showed that deletion of the hydrophillic cytosolic domain affects translocation of the NEP polypeptide chain. Substitution of a positively charged Lys residue for the Glu residue at the NH2-terminus of the deletion mutant only partly restored translocation of the polypeptide chain. Furthermore, carbonate extraction and trypsin digestion of the microsomal membranes indicated that the deletion mutants are inserted in the microsomal membranes as type III membrane proteins with their COOH-terminal domain exposed on the exterior of the microsomes. Thus, efficient translocation is dependent on the presence of a charged cytoplasmic domain.
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PMID:Translocation of neutral endopeptidase 24.11 mutants with deletions of the NH2-terminal cytosolic domain. 784 Sep 37

Mediastinal large B cell lymphomas are uncommon neoplasms that are thought to originate from thymic B cells. An unusual feature of these neoplasms is that they often lack surface immunoglobulin (Ig), a molecule ubiquitously expressed by most mature B cells. In the present study we have analyzed 12 cases of mediastinal large B cell lymphoma for the expression of the mb-1/CD79a polypeptide. This is a component, together with B29/CD79b, of a heterodimer that is associated with surface Ig on normal B cells. Our aim was to see whether loss of Ig in this type of lymphoma is associated with loss of the accompanying CD79a molecule. We have also evaluated 128 B cell lymphomas of other categories to see whether any of them show discordance between mb-1 and Ig expression and analyzed 30 T cell lymphomas as Ig-negative controls. We found that 5 of the 7 mediastinal large B cell lymphomas with interpretable staining results for both mb-1 and Ig, lack Ig but expressed CD79a (mb-1). This phenotype was very rare in other categories of B cell lymphoma, being found among 110 cases in only 5 cases that were all follicular lymphoma. The remaining 105 B cell lymphomas displayed mb-1+/Ig+ phenotype. All 30 T cell lymphomas were mb-1 negative. We conclude that discordant mb-1/Ig expression occurs commonly in mediastinal large B cell lymphomas. In addition, the finding that 11 of 12 of these neoplasms express a phenotype (CD10-, CD19+, CD20+, CD21-, CD22+, CD23-/+) that is very similar to that described for thymic medullary B cells reinforces the idea that most mediastinal large B cell lymphomas are of thymic B cell origin. The correlation between mb-1 and Ig staining patterns in B cell lymphomas of other categories reveals that in the majority (90%), expression of the antigen receptor complex parallels that of mature B cells. These data therefore confirm that the expression of the mb-1 protein provides independent strong evidence for the B lineage of lymphomas and may be used for their routine phenotypic characterization.
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PMID:Discordant expression of immunoglobulin and its associated molecule mb-1/CD79a is frequently found in mediastinal large B cell lymphomas. 788 54

Three subunits, Ac115, Ac39, and the proteolipid, were positively identified in the membrane sectors of V-ATPases from different sources. We searched for organelle-specific protein in purified preparations of V-ATPase from bovine chromaffin granules. A diffused protein band at a position of about 45 kDa was identified in SDS-polyacrylamide gels of the above preparation. Following digestion with endopeptidase Glu-C (V-8), a polypeptide of about 10 kDa was isolated and subjected to amino acid sequencing. Hence, the cDNA encoding the protein Ac45 was cloned from a bovine adrenal medulla library. The cDNA sequence contains an open reading frame encoding a protein of 468 amino acids with a calculated molecular mass of 51,786 daltons. A potential signal sequence comprised of the first 35 amino acids and a potential transmembrane domain at the C terminus of the protein were identified. There exist seven potential glycosylation sites between the aforementioned protein motifs. Experiments with a specific antibody against Ac45 demonstrated that it is copurifying with the V-ATPase from chromaffin granules. Immunological cross-reactivity was observed with purified V-ATPase from bovine kidney microsomes but not from plasma membranes of epithelial cells. Cell-free expression of the protein from synthetic mRNA produced a single protein band at about 50 kDa on SDS gels. Upon inclusion of dog pancreas microsomes in the reaction mixture, a slow migrating band sensitive to peptide:N-glycosidase F was observed.
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PMID:A novel accessory subunit for vacuolar H(+)-ATPase from chromaffin granules. 792 63

The interaction of different polypeptide growth factors with their cell surface receptors, which are typically rich in cysteine, is modulated by sulfhydryl (SH) reagents. Because the extracellular domain of both human TNFRs are likewise rich in cysteine residues, we examined the effect of SH reagents on these receptors in human histiocytic lymphoma U937 cells, which express both p60 and p80 forms. Iodoacetamide induced down-regulation of cell surface TNFRs in a dose- and time-dependent manner. Down-regulation was complete at 10 mM IAA for 1 h at 37 degrees C. The decrease was minimal at 4 degrees C. The down-modulation was also observed with other cell-permeable and -impermeable thiol reagents. The expression of other cell surface molecules such as CD3, CD11b, CD23, and CD25 were not affected. IAA induced the down-regulation of TNFR on cells of both epithelial and myeloid origin. With the use of receptor-specific Abs, we found that the kinetics of down-modulation of the p60 and p80 receptors by IAA were very similar. We also found that down-modulation was not a result of internalization but rather of the shedding of the receptors, as ascertained by receptor-ligand cross-linking analysis, immunoassay, Western blot analysis, and ligand-blot analysis. When TNFRs with a molecular mass of 80 kDa disappeared from the cell surface, a 42-kDa polypeptide appeared in the medium. Thus, we demonstrate that SH reagents down-regulate TNFRs by inducing shedding of both receptors from the cell surface, most likely by activating an endopeptidase that cleaves the receptor.
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PMID:Role of sulfhydryl groups in induction of cell surface down-modulation and shedding of extracellular domain of human TNF receptors in human histiocytic lymphoma U937 cells. 793 May 91

The C-terminal part of ligand filled porcine estradiol receptor extending from H267 to I595 was isolated by adsorption to the monoclonal antibody 13H2, subjected to cleavage by CNBr, o-iodosobenzoic acid and endopeptidase Lys-C as well as other proteases, both in the native and the denatured state. The overlapping peptides produced were separated by reverse phase HPLC and sequenced by Edman degradation, lacking T570-M581 in domain F. We found no evidence of post-translational modification; the native fragment is not glycosylated and the tyrosyl residues in domain E (aa 328, 331, 459, 526, 537) and F (aa 582, 583) are not phosphorylated. In addition, all serine and threonine PTH derivatives were obtained in normal yields. The amino acid sequence of the fragment corresponds in full with that derived from the cDNA. The complete cDNA-derived sequence codes for a polypeptide of 595 amino acids with a calculated mass of 66,357 Da. The high degree of homology between species in domains C and E is shared by the porcine receptor.
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PMID:The C-terminal half of the porcine estradiol receptor contains no post-translational modification: determination of the primary structure. 798 44


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