Linked Life Data

15044477

Source:http://linkedlifedata.com/resource/pubmed/id/15044477

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
21
pubmed:dateCreated
2004-5-17
pubmed:abstractText
We have shown previously that the 134-residue endonuclease domain of the bacterial cytotoxin colicin E9 (E9 DNase) forms channels in planar lipid bilayers (Mosbahi, K., Lemaître, C., Keeble, A. H., Mobasheri, H., Morel, B., James, R., Moore, G. R., Lea, E. J., and Kleanthous, C. (2002) Nat. Struct. Biol. 9, 476-484). It was proposed that the E9 DNase mediates its own translocation across the cytoplasmic membrane and that the formation of ion channels is essential to this process. Here we describe changes to the structure and stability of the E9 DNase that accompany interaction of the protein with phospholipid vesicles. Formation of the protein-lipid complex at pH 7.5 resulted in a red-shift of the intrinsic protein fluorescence emission maximum (lambda(max)) from 333 to 346 nm. At pH 4.0, where the E9 DNase lacks tertiary structure but retains secondary structure, DOPG induced a blue-shift in lambda(max), from 354 to 342 nm. Changes in lambda(max) were specific for anionic phospholipid vesicles at both pHs, suggesting electrostatics play a role in this association. The effects of phospholipid were negated by Im9 binding, the high affinity, acidic, exosite inhibitor protein, but not by zinc, which binds at the active site. Fluorescence-quenching experiments further demonstrated that similar protein-phospholipid complexes are formed regardless of whether the E9 DNase is initially in its native conformation. Consistent with these observations, chemical and thermal denaturation data as well as proteolytic susceptibility experiments showed that association with negatively charged phospholipids destabilize the E9 DNase. We suggest that formation of a destabilizing protein-lipid complex pre-empts channel formation by the E9 DNase and constitutes the initial step in its translocation across the Escherichia coli inner membrane.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
May
pubmed:issn
0021-9258
pubmed:author
pubmed:issnType
Print
pubmed:day
21
pubmed:volume
279
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
22145-51
pubmed:dateRevised
2008-11-21
pubmed:meshHeading
pubmed-meshheading:15044477-Acrylamide, pubmed-meshheading:15044477-Circular Dichroism, pubmed-meshheading:15044477-Colicins, pubmed-meshheading:15044477-Cytoplasm, pubmed-meshheading:15044477-Deoxyribonucleases, pubmed-meshheading:15044477-Disulfides, pubmed-meshheading:15044477-Dose-Response Relationship, Drug, pubmed-meshheading:15044477-Endonucleases, pubmed-meshheading:15044477-Escherichia coli, pubmed-meshheading:15044477-Hydrogen-Ion Concentration, pubmed-meshheading:15044477-Ions, pubmed-meshheading:15044477-Ligands, pubmed-meshheading:15044477-Lipid Bilayers, pubmed-meshheading:15044477-Lipids, pubmed-meshheading:15044477-Phospholipids, pubmed-meshheading:15044477-Protein Binding, pubmed-meshheading:15044477-Protein Conformation, pubmed-meshheading:15044477-Protein Structure, Tertiary, pubmed-meshheading:15044477-Protein Transport, pubmed-meshheading:15044477-Spectrometry, Fluorescence, pubmed-meshheading:15044477-Static Electricity, pubmed-meshheading:15044477-Temperature, pubmed-meshheading:15044477-Urea
pubmed:year
2004
pubmed:articleTitle
Destabilization of the colicin E9 Endonuclease domain by interaction with negatively charged phospholipids: implications for colicin translocation into bacteria.
pubmed:affiliation
Department of Biology, University of York, York YO10 5YW, United Kingdom.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't