Source:http://linkedlifedata.com/resource/pubmed/id/12452439
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rdf:type | |
lifeskim:mentions | |
pubmed:dateCreated |
2002-11-27
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pubmed:abstractText |
The vitamin D-binding protein (DBP) binds to monomeric actin with high affinity. The variation in DBP isoforms is due to genetic polymorphism and varying glycosylation. To obtain a homogeneous preparation, the cDNA for human DBP and truncations thereof were cloned and various systems were applied for heterologous bacterial and yeast expression. The full-length protein and the N- and C-terminal halves of DBP remained insoluble probably because the protein did not fold to its native three-dimensional structure due to formation of accidental intra- and inter-molecular disulfide bonds during expression in bacteria or yeast. This problem was overcome by cloning of a C-terminal fragment comprising residues 369 to 435 that did not contain disulfide bonds and was completely soluble. Binding of the C-terminal fragment to monomeric actin was demonstrated by comigration with actin during native polyacrylamide gel electrophoresis and surface plasmon resonance, however, at considerably lower affinity than full-length DBP. This suggests that in addition to the C-terminal amino acid sequence other parts (amino acid residues or sugar moieties) of DBP participate in actin binding. The C-terminal fragment was found to inhibit denaturation of actin and to decrease the rate of actin polymerisation both at the barbed and at the pointed end in a concentration-dependent manner. According to a quantitative analysis of the polymerisation kinetics, association of actin monomers to nucleate filaments was not prevented by binding of the C-terminal fragment to actin. These data suggest that the sites on the surface of actin that are involved in actin nucleation and elongation are different.
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pubmed:language |
eng
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pubmed:citationSubset |
IM
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pubmed:chemical |
http://linkedlifedata.com/resource/pubmed/chemical/Actins,
http://linkedlifedata.com/resource/pubmed/chemical/DNA Primers,
http://linkedlifedata.com/resource/pubmed/chemical/Peptide Fragments,
http://linkedlifedata.com/resource/pubmed/chemical/Polymers,
http://linkedlifedata.com/resource/pubmed/chemical/Vitamin D-Binding Protein
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pubmed:status |
MEDLINE
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pubmed:author | |
pubmed:owner |
NLM
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pubmed:authorsComplete |
Y
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pubmed:pagination |
1621-31
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pubmed:dateRevised |
2004-11-17
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pubmed:articleTitle |
Binding of a C-terminal fragment (residues 369 to 435) of vitamin D-binding protein to actin.
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pubmed:affiliation |
Department of Anatomy and Embryology, Ruhr-University Bochum, D-44780 Bochum, Germany.
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