Linked Life Data

2833171

Source:http://linkedlifedata.com/resource/pubmed/id/2833171

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
1
pubmed:dateCreated
1988-5-10
pubmed:abstractText
Soluble extracts of cultured cells (BHK 21/C13) degraded a variety of exogenous proteins to acid-soluble peptides at pH 8.0. ATP stimulated this proteolytic activity up to 10-fold. The ATP effect was dependent on Mg2+ and was not elicited by nonhydrolyzable analogs of ATP. After the extract was fractionated on DEAE-cellulose, ATP-stimulated protease activity was in the fraction that bound to the resin and eluted in buffer containing 0.4 M NaCl. This activity had characteristics that were indistinguishable from those of the unfractionated extract but the degree of ATP stimulation was two- to three-fold lower. Although no protease activity was detected in the unbound fraction, reconstitution of this material with the bound fraction enhanced the ATP stimulation up to twofold. The component responsible for the enhancement of the ATP stimulation had properties similar to ubiquitin and purified ubiquitin enhanced the ATP-stimulated protease activity in the fractionated extract. Substrates whose amino groups were almost completely blocked by various chemical modifications were still degraded in an ATP-stimulated fashion, but the degradation of these substrates was not affected by ubiquitin. The protease activity isolated by ion-exchange chromatography was fractionated further by gel filtration chromatography on Sephacryl S-300. ATP-stimulated protease activity eluted with an apparent molecular weight of 750,000. Protease activity was enhanced up to eightfold by Mg2+-ATP but was not increased further by ubiquitin. An activity that hydrolyzed the synthetic peptide Z-Val-Leu-Arg-MNA coeluted with ATP-stimulated protease activity, but peptide hydrolysis was not affected by ATP. These and other catalytic and biochemical characteristics suggested that the protease might be related to the high-molecular-weight protease, macropain, recently purified by us from human erythrocytes (M. J. McGuire and G. N. DeMartino Biochim. Biophys. Acta (1986) 873, 279-289). Antibodies raised against macropain specifically reacted with proteins characteristic of macropain in the column fractions containing ATP-stimulated protease activity. These antibodies also specifically immunoprecipitated 70-100% of the ATP-stimulated protease activity as well as Z-Val-Leu-Arg-MNA hydrolyzing activity. Thus BHK cell extracts appear to contain both ubiquitin-mediated and ubiquitin-independent pathways for the ATP-stimulated degradation of proteins. Furthermore, at least one of these pathways appears to involve a high-molecular-weight, ATP-stimulated protease related to macropain.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Apr
pubmed:issn
0003-9861
pubmed:author
pubmed:issnType
Print
pubmed:volume
262
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
273-85
pubmed:dateRevised
2007-11-14
pubmed:meshHeading
pubmed:year
1988
pubmed:articleTitle
ATP-stimulated proteolysis in soluble extracts of BHK 21/C13 cells. Evidence for multiple pathways and a role for an enzyme related to the high-molecular-weight protease, macropain.
pubmed:affiliation
Department of Physiology, University of Texas Health Science Center, Dallas 75235-9040.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S.