Linked Life Data

12930827

Source:http://linkedlifedata.com/resource/pubmed/id/12930827

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
44
pubmed:dateCreated
2003-10-27
pubmed:abstractText
We have studied Hansenula polymorpha Pex5p and Pex8p using fluorescence correlation spectroscopy (FCS). Pex5p is the Peroxisomal Targeting Signal 1 (PTS1) receptor and Pex8p is an intraperoxisomal protein. Both proteins are essential for PTS1 protein import and have been shown to physically interact. We used FCS to analyze the molecular role of this interaction. FCS is a very sensitive technique that allows analysis of dynamic processes of fluorescently marked molecules at equilibrium in a very tiny volume. We used this technique to determine the oligomeric state of both peroxins and to analyze binding of Pex5p to PTS1 peptides and Pex8p. HpPex5p and HpPex8p were overproduced in Escherichia coli, purified by affinity chromatography, and, when required, labeled with the fluorescent dye Alexa Fluor 488. FCS measurements revealed that the oligomeric state of HpPex5p varied, ranging from monomers at slightly acidic pH to tetramers at neutral pH. HpPex8p formed monomers at all pH values tested. Using fluorescein-labeled PTS1 peptide and unlabeled HpPex5p, we established that PTS1 peptide only bound to tetrameric HpPex5p. Upon addition of HpPex8p, a heterodimeric complex was formed consisting of one HpPex8p and one HpPex5p molecule. This process was paralleled by dissociation of PTS1 peptide from HpPex5p, indicating that Pex8p may play an important role in cargo release from the PTS1 receptor. Our data show that FCS is a powerful technique to explore dynamic physical interactions that occur between peroxins during peroxisomal matrix protein import.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Oct
pubmed:issn
0021-9258
pubmed:author
pubmed:issnType
Print
pubmed:day
31
pubmed:volume
278
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
43340-5
pubmed:dateRevised
2006-11-15
pubmed:meshHeading
pubmed-meshheading:12930827-Diffusion, pubmed-meshheading:12930827-Dimerization, pubmed-meshheading:12930827-Fluorescent Dyes, pubmed-meshheading:12930827-Hydrogen-Ion Concentration, pubmed-meshheading:12930827-Ligands, pubmed-meshheading:12930827-Membrane Transport Proteins, pubmed-meshheading:12930827-Microscopy, Confocal, pubmed-meshheading:12930827-Models, Biological, pubmed-meshheading:12930827-Models, Statistical, pubmed-meshheading:12930827-Peptides, pubmed-meshheading:12930827-Pichia, pubmed-meshheading:12930827-Plasmids, pubmed-meshheading:12930827-Protein Binding, pubmed-meshheading:12930827-Protein Transport, pubmed-meshheading:12930827-Receptors, Cytoplasmic and Nuclear, pubmed-meshheading:12930827-Saccharomyces cerevisiae Proteins, pubmed-meshheading:12930827-Spectrometry, Fluorescence
pubmed:year
2003
pubmed:articleTitle
Physical interactions of the peroxisomal targeting signal 1 receptor pex5p, studied by fluorescence correlation spectroscopy.
pubmed:affiliation
Eukaryotic Microbiology, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, P.O. Box 14, 9750AA, Haren, The Netherlands.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't