| Expression of rat kallikrein and epithelial polarity in transfected Madin-Darby canine kidney cells. | |
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MedLine Citation:
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PMID: 7490145 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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Many properties of urinary kallikrein are well characterized, but the intracellular processing of prokallikrein and release by kidney cells have yet to be clarified. We report here on the synthesis of prokallikrein in Madin-Darby canine kidney (MDCK) cells transfected with rat submaxillary gland kallikrein cDNA and on its activation by MDCK cells and by an enriched liver Golgi membrane preparation. Transfected MDCK cells secreted only prokallikrein at both the apical and basolateral sides in about a 4:1 ratio, but cells transfected with kallikrein cDNA in reverse orientation or untreated cells released only traces of the enzyme. Prokallikrein, in culture medium or in homogenized MDCK cells, was fully activated by trypsin but activated only to 44% by thermolysin. Prokallikrein was synthesized and released into the medium at a high rate: the enzyme secreted by 5 x 10(6) cells in 24 hours cleaved 46 nmol/min D-Val-Leu-Arg-7-amino-4-methylcoumarin and liberated 63 ng/min bradykinin after activation. Immunocytology indicated the association of prokallikrein with the Golgi apparatus in the transfected cells. Antiserum to rat urinary kallikrein detected a single band in a Western blot of conditioned medium and also immunoprecipitated the enzyme. Aprotinin inhibited activated prokallikrein. Although MDCK cells released prokallikrein, their homogenates activated prokallikrein at both pH 5.5 and 7.5. Prokallikrein was also activated by a highly enriched liver Golgi membrane fraction and by an endoplasmic reticulum preparation, but the Golgi preparation was 38-fold more active. The activation was blocked significantly by inhibitors of serine proteases and less by cysteine protease inhibitors.(ABSTRACT TRUNCATED AT 250 WORDS) |
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Authors:
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M Abe; F Nakamura; F Tan; P A Deddish; K J Colley; R P Becker; R A Skidgel; E G Erdös |
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Publication Detail:
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Type: Comparative Study; Journal Article; Research Support, U.S. Gov't, P.H.S. |
Journal Detail:
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Title: Hypertension Volume: 26 ISSN: 0194-911X ISO Abbreviation: Hypertension Publication Date: 1995 Dec |
Date Detail:
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Created Date: 1996-01-04 Completed Date: 1996-01-04 Revised Date: 2007-11-14 |
Medline Journal Info:
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Nlm Unique ID: 7906255 Medline TA: Hypertension Country: UNITED STATES |
Other Details:
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Languages: eng Pagination: 891-8 Citation Subset: IM |
Affiliation:
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Department of Pharmacology, University of Illinois College of Medicine at Chicago 60612, USA. |
Export Citation:
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APA/MLA Format Download EndNote Download BibTex |
| MeSH Terms | |
Descriptor/Qualifier:
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Animals Blotting, Western Cell Line Culture Media DNA, Complementary / genetics Dogs Endoplasmic Reticulum / enzymology Enzyme Activation Enzyme Precursors / biosynthesis*, secretion Golgi Apparatus / enzymology Immunohistochemistry Kallikreins / genetics*, metabolism* Kidney / enzymology*, secretion Precipitin Tests Rats Recombinant Proteins / genetics Submandibular Gland / enzymology Time Factors Transfection* / genetics |
| Grant Support | |
ID/Acronym/Agency:
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HL-36081/HL/NHLBI NIH HHS; HL-36473/HL/NHLBI NIH HHS |
| Chemical | |
Reg. No./Substance:
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0/Culture Media; 0/DNA, Complementary; 0/Enzyme Precursors; 0/Recombinant Proteins; EC 3.4.21.-/Kallikreins |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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