Linked Life Data

14660572

Source:http://linkedlifedata.com/resource/pubmed/id/14660572

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
8
pubmed:dateCreated
2004-2-17
pubmed:abstractText
Ficolin is a plasma lectin, consisting of a short N-terminal multimerization domain, a middle collagen domain, and a C-terminal fibrinogen-like domain. The collagen domains assemble the subunits into trimers, and the N-terminal domain assembles four trimers into 12-mers. Two cysteine residues in the N-terminal domain are thought to mediate multimerization by disulfide bonding. We have generated three mutants of ficolin alpha in which the N-terminal cysteines were substituted by serines (Cys4, Cys24, and Cys4/Cys24). The N-terminal cysteine mutants were produced in a mammalian cell expression system, purified by affinity chromatography, and analyzed under nondenaturing conditions to resolve the multimer structure of the native protein and under denaturing conditions to resolve the disulfide-linked structure. Glycerol gradient sedimentation and electron microscopy in nondenaturing conditions showed that plasma and recombinant wild-type protein formed 12-mers. The Cys4 mutant also formed 12-mers, but Cys24 and Cys4/Cys24 mutants formed only trimers. This means that protein interfaces containing Cys4 are stable as noncovalent protein-protein interactions and do not require disulfides, whereas those containing Cys24-Cys24 require the disulfides for stability. Proteins were also analyzed by nonreducing SDS-PAGE to show the covalent structure under denaturing conditions. Wild-type ficolin was covalently linked into 12-mers, whereas elimination of either Cys4 or Cys24 gave dimers and monomers. We present a model in which symmetric Cys24-Cys24 disulfide bonds between trimers are the basis for multimerization. The model may also be relevant to collectin multimers.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Feb
pubmed:issn
0021-9258
pubmed:author
pubmed:issnType
Print
pubmed:day
20
pubmed:volume
279
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
6534-9
pubmed:dateRevised
2007-11-14
pubmed:meshHeading
pubmed-meshheading:14660572-Amino Acid Sequence, pubmed-meshheading:14660572-Animals, pubmed-meshheading:14660572-Blotting, Western, pubmed-meshheading:14660572-CHO Cells, pubmed-meshheading:14660572-Carrier Proteins, pubmed-meshheading:14660572-Chromatography, Affinity, pubmed-meshheading:14660572-Cricetinae, pubmed-meshheading:14660572-Culture Media, Conditioned, pubmed-meshheading:14660572-Cysteine, pubmed-meshheading:14660572-Dimerization, pubmed-meshheading:14660572-Disulfides, pubmed-meshheading:14660572-Electrophoresis, Polyacrylamide Gel, pubmed-meshheading:14660572-Glycerol, pubmed-meshheading:14660572-Lectins, pubmed-meshheading:14660572-Microscopy, Electron, pubmed-meshheading:14660572-Models, Molecular, pubmed-meshheading:14660572-Molecular Sequence Data, pubmed-meshheading:14660572-Mutation, pubmed-meshheading:14660572-Oxygen, pubmed-meshheading:14660572-Protein Conformation, pubmed-meshheading:14660572-Protein Structure, Tertiary, pubmed-meshheading:14660572-Recombinant Proteins, pubmed-meshheading:14660572-Sequence Homology, Amino Acid, pubmed-meshheading:14660572-Trypsin
pubmed:year
2004
pubmed:articleTitle
The disulfide bonding pattern in ficolin multimers.
pubmed:affiliation
Department of Cell Biology, Duke University Medical Center, Durham, North Carolina 27710, USA.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S.