Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.23.5 (cathepsin D)
 4,130 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Thiol and aspartyl proteolytic activities in isolated secretory vesicles of neural (NL) and intermediate (IL) lobes of bovine pituitary were characterized with heterologous enkephalin and tachykinin precursor substrates, 35S-(Met)-preproenkephalin and 35S-(Met)-beta-preprotachykinin. IL and NL secretory vesicles contained thiol-dependent proteolytic activity that cleaved the enkephalin precursor with a pH optimum of 4.5; this activity resembled a novel "prohormone thiol protease' previously purified and characterized from adrenal medulla chromaffin granules. IL and NL vesicles also demonstrated aspartyl proteolytic activity with acidic pH optimum, as shown by pepstatin A inhibition of tachykinin and enkephalin precursor cleaving activity. This activity may be related to a previously characterized chromaffin granule aspartyl protease (CGAP) related to cathepsin D (2), as indicated by the presence of immunoreactive CGAP in NL secretory vesicles by anti-CGAP immunoblots. These results show that pituitary secretory vesicles, like chromaffin granules, may contain similar thiol-dependent and aspartyl proteolytic activities.
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PMID:Thiol and aspartyl proteolytic activities in secretory vesicles of bovine pituitary. 155 May 54

Purification and potential tachykinin and enkephalin precursor cleaving enzymes from bovine chromaffin granules was undertaken using as substrates the model precursors 35S-(Met)-beta-preprotachykinin [35S-(Met)-beta-PPT] and 35S-(Met)-preproenkephalin [35S-(Met)-PPE]. Purification by concanavalin A-Sepharose, Sephacryl S200, and chromatofocusing resulted in a chromaffin granule aspartyl protease (CGAP) that preferred the tachykinin over the enkephalin precursor. CGAP was composed of 47-, 30-, and 16.5-kDa polypeptides migrating as a single band in a nondenaturing electrophoretic gel system, and coeluting with an apparent molecular mass of 45-55 kDa by size-exclusion chromatography. These results suggest that two forms exist: a single 47-kDa polypeptide and a complex of 30 + 16.5-kDa-associated subunits. CGAP was optimally active at pH 5.0-5.5, indicating that it would be active within the acidic intragranular environment. Cleavage at basic residues was suggested by HPLC and HVE identification of 35S-(Met)-NKA-Gly-Lys as the major acid-soluble product generated from 35S-(Met)-beta-PPT. Neuropeptide K was cleaved at a Lys-Arg basic residue site, as determined by identification of proteolytic products by microsequencing and amino acid composition analyses. Structural studies showed that the three CGAP polypeptides were similar to bovine cathepsin D in NH2-terminal sequences and amino acid compositions, indicating that CGAP appears to be a cathepsin D-related protease or cathepsin D itself. The 47- and 16.5-kDa polypeptides of CGAP possessed identical NH2-terminal sequences, suggesting that the 16.5-kDa polypeptide may be derived from the 47-kDa form by proteolysis.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Purification and characterization of a cathepsin D protease from bovine chromaffin granules. 156 70

Proteolytic processing of neuropeptide precursors is required for production of active neurotransmitters and hormones. In this study, a chromaffin granule (CG) aspartic proteinase of 70 kDa was found to contribute to enkephalin precursor cleaving activity, as assayed with recombinant ([35S]Met) preproenkephalin. The 70-kDa CG aspartic proteinase was purified by concanavalin A-Sepharose, Sephacryl S-200, and pepstatin A agarose affinity chromatography. The proteinase showed optimal activity at pH 5.5. It was potently inhibited by pepstatin A, a selective aspartic proteinase inhibitor, but not by inhibitors of serine, cysteine, or metalloproteinases. Lack of inhibition by Val-D-Leu-Pro-Phe-Val-D-Leu--an inhibitor of pepsin, cathepsin D, and cathepsin E--distinguishes the CG aspartic proteinases from classical members of the aspartic proteinase family. The CG aspartic proteinase cleaved recombinant proenkephalin between the Lys172-Arg173 pair located at the COOH-terminus of (Met)enkephalin-Arg6-Gly7-Leu8, as assessed by peptide microsequencing. The importance of full-length prohormone as substrate was demonstrated by the enzyme's ability to hydrolyze 35S-labeled proenkephalin and proopiomelanocortin and its inability to cleave tri- and tetrapeptide substrates containing dibasic or monobasic cleavage sites. In this study, results provide evidence for the role of an aspartic proteinase in proenkephalin and prohormone processing.
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PMID:Characteristics of the chromaffin granule aspartic proteinase involved in proenkephalin processing. 756 75