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| Predicate | Object |
|---|---|
| rdf:type | |
| lifeskim:mentions | |
| pubmed:issue |
2
|
| pubmed:dateCreated |
1995-11-22
|
| pubmed:abstractText |
The extracellular glycoprotein BM-40 consists of three domains, an acidic domain I, a follistatin (FS)-like domain II and a calcium-binding EC domain with an EF-hand related motif. BM-40 and several other related proteins (QR1, SC1/hevin, testican and tsc-36/FRP) are members of a novel modular protein family that share the FS domain followed by an EC domain. We have expressed this pair of FS and EC domains (mutant delta I) and the calcium-binding EC domain alone (mutant delta I, II) of human BM-40 as recombinant proteins in human 293 cells. Circular dichroism demonstrated that both mutants were obtained as folded proteins with a distinct three-dimensional conformation. In addition, mutant delta I, II could be readily crystallized and diffraction patterns with a resolution limit of 2.4 A resolution were obtained. Calcium binding to this fragment was ten times weaker (Kd = 0.8 microM) than for the wild-type protein. Identical reversible increases in alpha-helicity upon calcium binding were observed for the 150-residue long mutant delta I, II and for BM-40 (286 residues). A 26-residue synthetic peptide corresponding to the EF-hand related motif exhibited much weaker calcium binding. The apparent dissociation constant decreased with increasing peptide concentration (from Kd 2.4 mM at 1 microM, to Kd 0.3 mM at 100 microM peptide concentration) and calcium binding was accompanied by dimerization of the peptide. This suggests that for strong calcium binding the EF-hand related motif has to be embedded into a larger protein domain that can form an autonomously folding protein module. The EC domain was also shown by surface plasmon resonance assay to be responsible for calcium-dependent binding to collagen IV with an affinity (Kd = 19 microM) only sixfold lower than that of intact human BM-40.
|
| pubmed:language |
eng
|
| pubmed:journal | |
| pubmed:citationSubset |
IM
|
| pubmed:chemical |
http://linkedlifedata.com/resource/pubmed/chemical/Calcium,
http://linkedlifedata.com/resource/pubmed/chemical/Collagen,
http://linkedlifedata.com/resource/pubmed/chemical/DNA Primers,
http://linkedlifedata.com/resource/pubmed/chemical/Osteonectin,
http://linkedlifedata.com/resource/pubmed/chemical/Peptide Fragments,
http://linkedlifedata.com/resource/pubmed/chemical/Recombinant Proteins
|
| pubmed:status |
MEDLINE
|
| pubmed:month |
Oct
|
| pubmed:issn |
0022-2836
|
| pubmed:author | |
| pubmed:issnType |
Print
|
| pubmed:day |
20
|
| pubmed:volume |
253
|
| pubmed:owner |
NLM
|
| pubmed:authorsComplete |
Y
|
| pubmed:pagination |
347-57
|
| pubmed:dateRevised |
2006-11-15
|
| pubmed:meshHeading |
pubmed-meshheading:7563094-Amino Acid Sequence,
pubmed-meshheading:7563094-Base Sequence,
pubmed-meshheading:7563094-Binding Sites,
pubmed-meshheading:7563094-Calcium,
pubmed-meshheading:7563094-Cell Line,
pubmed-meshheading:7563094-Circular Dichroism,
pubmed-meshheading:7563094-Collagen,
pubmed-meshheading:7563094-Crystallography, X-Ray,
pubmed-meshheading:7563094-DNA Primers,
pubmed-meshheading:7563094-Humans,
pubmed-meshheading:7563094-Kidney,
pubmed-meshheading:7563094-Kinetics,
pubmed-meshheading:7563094-Molecular Sequence Data,
pubmed-meshheading:7563094-Mutagenesis, Site-Directed,
pubmed-meshheading:7563094-Osteonectin,
pubmed-meshheading:7563094-Peptide Fragments,
pubmed-meshheading:7563094-Polymerase Chain Reaction,
pubmed-meshheading:7563094-Protein Folding,
pubmed-meshheading:7563094-Protein Structure, Secondary,
pubmed-meshheading:7563094-Recombinant Proteins,
pubmed-meshheading:7563094-Restriction Mapping,
pubmed-meshheading:7563094-Transfection
|
| pubmed:year |
1995
|
| pubmed:articleTitle |
The C-terminal portion of BM-40 (SPARC/osteonectin) is an autonomously folding and crystallisable domain that binds calcium and collagen IV.
|
| pubmed:affiliation |
Abt. Biophysikalische Chemie, Biozentrum University, Basel, Switzerland.
|
| pubmed:publicationType |
Journal Article,
Comparative Study,
Research Support, Non-U.S. Gov't
|