HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) All Versions of this Article: 292/5/L1280    most recent 00140.2006v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in Web of Science Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Web of Science (6) Citing Articles via Google Scholar Google Scholar Articles by Sixt, S. U. Articles by Peters, J. Search for Related Content PubMed PubMed Citation Articles by Sixt, S. U. Articles by Peters, J.

Extracellular proteasome in the human alveolar space: a new housekeeping enzyme?

¹Klinik für Anästhesiologie und Intensivmedizin, Universitätsklinikum Essen, and ²Institut für Physiologische Chemie II, Universität Duisburg-Essen, Essen, Germany

Submitted 13 April 2006 ; accepted in final form 1 January 2007

We hypothesized that 20S proteasome is present and functional in the extracellular alveolar space in humans. Proteasomal activity was measured in bronchoalveolar lavage (BAL) supernatant from eight humans using specific proteasomal fluorogenic substrates and I¹²⁵-albumin with and without specific proteasome inhibitors. Furthermore, gelfiltration, Western blot technique, and mass spectrometry were applied for proteasome characterization. All proteasomal fluorogenic substrates were hydrolyzed by BAL supernatant, with hydrolysis inhibited by epoxomicin (P = 0.024) and other proteasome inhibitors as well. E64, a lysosomal inhibitor, did not inhibit enzyme activity. The majority of proteolytic activity was detected in BAL supernatant rather than in the cell pellet. No correlation was found between proteasomal hydrolysis in BAL supernatant and lactate dehydrogenase activity, the total cell count in the cell pellet, and the fraction of avital cells in the cell pellet, ruling out cell lysis as a major source of proteasomal activity. Gelfiltration revealed hydrolyzing activity in the supernatant at 660 kDa and proteasome core proteins after analysis by ESI-QqTOF mass spectrometry. Furthermore, Western blots using a polyclonal antibody against proteasomal -/-subunits detected proteasomal proteins in the typical 20- to 30-kDa range in BAL supernatant. Incubation of BAL supernatant with I¹²⁵-albumin showed a high mean cleavage rate (101.8 µg/ml x h lavage ± 46 SD) that was inhibited by epoxomicin (P = 0.013) and was ATP and ubiquitin independent. We identified for the first time extracellular, biologically active, ATP- and ubiquitin-independent 20S proteasome in the human alveolar space, with a high albumin cleavage rate. Possibly, the proteasome assists in maintenance of a low intra-alveolar oncotic pressure and/or alveolar protein degradation.

albumin; bronchoalveolar lavage; circulating proteasome; fluorogenic peptides; alveolar protein degradation; lung proteins

This article has been cited by other articles:

				S. U. Sixt, M. Adamzik, D. Spyrka, B. Saul, J. Hakenbeck, J. Wohlschlaeger, U. Costabel, A. Kloss, J. Giesebrecht, B. Dahlmann, et al. Alveolar Extracellular 20S Proteasome in Patients with Acute Respiratory Distress Syndrome Am. J. Respir. Crit. Care Med., June 15, 2009; 179(12): 1098 - 1106. [Abstract] [Full Text] [PDF]

				W. Huang, S. Y. Eum, I. E Andras, B. Hennig, and M. Toborek PPAR{alpha} and PPAR{gamma} attenuate HIV-induced dysregulation of tight junction proteins by modulations of matrix metalloproteinase and proteasome activities FASEB J, May 1, 2009; 23(5): 1596 - 1606. [Abstract] [Full Text] [PDF]

				G. A. Cornwall New insights into epididymal biology and function Hum. Reprod. Update, March 1, 2009; 15(2): 213 - 227. [Abstract] [Full Text] [PDF]