Linked Life Data

1535223

Source:http://linkedlifedata.com/resource/pubmed/id/1535223

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
3
pubmed:dateCreated
1992-7-28
pubmed:abstractText
The dissociation of mitochondrial F1-ATPase with 3 M LiCl at 0 degrees C, followed by reconstitution, has been analysed. FPLC over a gel filtration column in the dissociation buffer revealed the presence of two protein moieties, an alpha 3 gamma delta epsilon complex and single beta-subunits. When the dissociation and chromatography is performed at pH 6.2, the former protein moiety still contains some adenine nucleotides. Reconstitution of the dissociated complex is not possible any more after FPLC, probably due to the loss of residual adenine nucleotides. After a single column centrifugation step one nucleotide per F1 still remains bound. For reconstitution, additional ATP, or a suitable analog, is required. 2-Azido-ATP, but not 8-azido-ATP or ITP, can replace ATP during the reconstitution. F1, reconstituted in the presence of 2-azido-ATP, contains three tightly bound nucleotides, similar to freshly isolated F1, of which in this case one is an adenine nucleotide and two are azido-adenine nucleotides. One of the latter can be rapidly exchanged and is bound to a catalytic site. Covalent binding (at a beta-subunit) of the other tightly bound 2-azido-ATP by ultraviolet illumination does not result in inhibition of the enzyme. Digestion of F1 with trypsin, followed by HPLC, showed that the label is not bound to the fragment containing Tyr-368, nor to the fragment containing Tyr-345. This result was confirmed by CNBr digestion, followed by SDS-urea PAGE. We conclude that during dissociation of F1 one tightly bound nucleotide (ADP) remains bound at an alpha/beta interface site and that for reconstitution binding of ATP to a (non-catalytic) beta-site is required. The conformation of this site differs from that of the two catalytic beta-sites.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Jun
pubmed:issn
0006-3002
pubmed:author
pubmed:issnType
Print
pubmed:day
19
pubmed:volume
1100
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
267-77
pubmed:dateRevised
2006-11-15
pubmed:meshHeading
pubmed:year
1992
pubmed:articleTitle
Characteristics of the non-exchangeable nucleotide binding sites of mitochondrial F1 revealed by dissociation and reconstitution with 2-azido-ATP.
pubmed:affiliation
E.C. Slater Institute for Biochemical Research, University of Amsterdam, Netherlands.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't