GONUTS has been updated to MW1.31 Most things seem to be working but be sure to report problems.

Have any questions? Please email us at ecoliwiki@gmail.com

PMID:8037676

From GONUTS
Jump to: navigation, search
Citation

McGregor, A, Blanchard, AD, Rowe, AJ and Critchley, DR (1994) Identification of the vinculin-binding site in the cytoskeletal protein alpha-actinin. Biochem. J. 301 ( Pt 1):225-33

Abstract

Using low-speed sedimentation equilibrium we have established that vinculin binds to alpha-actinin with a Kd of 1.3 x 10(-5) M. Electron microscopy of negatively stained preparations of vinculin revealed spherical particles (diameter 11.2 nm; S.D. 1.7 nm, n = 21), whereas alpha-actinin appeared as a rod-shaped particle (length 33 nm; S.D. 3.3 nm, n = 23). Mixtures of the two proteins contained both 'lollipop'- and 'dumbell'-shaped particles which we interpret as either one or two spherical vinculin molecules associated with the ends of the alpha-actinin rod. We have further defined the vinculin-binding site in alpha-actinin using 125I-vinculin and a gel-blot assay in which proteolytic fragments of alpha-actinin and fragments of alpha-actinin expressed in Escherichia coli were resolved by SDS/PAGE and blotted to nitrocellulose. 125I-vinculin bound to polypeptides derived from the spectrin-like repeat region of alpha-actinin, but did not bind to the actin-binding domain. Binding was inhibited by a 100-fold molar excess of unlabelled vinculin. Using a series of glutathione S-transferase fusion proteins we have mapped the vinculin-binding site to a region toward the C-terminal end of the molecule (alpha-actinin residues 713-749). 125I-vinculin also bound to fusion proteins containing this sequence which had been immobilized on glutathione-agarose beads. The vinculin-binding site is localized in a highly conserved region of the molecule close to the first of two EF-hand calcium-binding motifs.

Links

PubMed PMC1137166

Keywords

Actinin/genetics; Actinin/metabolism; Actinin/ultrastructure; Animals; Binding Sites; Chickens; Escherichia coli/genetics; Kinetics; Microscopy, Electron; Models, Biological; Protein Binding; Recombinant Fusion Proteins/genetics; Recombinant Fusion Proteins/metabolism; Recombinant Fusion Proteins/ultrastructure; Vinculin/metabolism; Vinculin/ultrastructure

Significance

Annotations

Gene product Qualifier GO Term Evidence Code with/from Aspect Extension Notes Status

CHICK:ZYX

enables

GO:0017166: vinculin binding

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete

CHICK:ZYX

enables

GO:0042803: protein homodimerization activity

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete

CHICK:ZYX

enables

GO:0051393: alpha-actinin binding

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete

CHICK:ZYX

enables

GO:0005515: protein binding

ECO:0000353: physical interaction evidence used in manual assertion

UniProtKB:P12003

F

Seeded From UniProt

complete

CHICK:VINC

enables

GO:0005515: protein binding

ECO:0000353: physical interaction evidence used in manual assertion

UniProtKB:P05094-2

F

Seeded From UniProt

complete

CHICK:ACTN1

part_of

GO:0005623: cell

ECO:0000314: direct assay evidence used in manual assertion

C

part_of:(UBERON:0005052)

Seeded From UniProt

complete

CHICK:ACTN1

enables

GO:0042803: protein homodimerization activity

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete

CHICK:ACTN1

enables

GO:0051393: alpha-actinin binding

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete

CHICK:ACTN1

enables

GO:0017166: vinculin binding

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete

CHICK:VINC

part_of

GO:0005623: cell

ECO:0000314: direct assay evidence used in manual assertion

C

part_of:(UBERON:0005052)

Seeded From UniProt

complete

CHICK:VINC

enables

GO:0042803: protein homodimerization activity

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete

CHICK:VINC

enables

GO:0051393: alpha-actinin binding

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete

CHICK:VINC

enables

GO:0017166: vinculin binding

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete

See also

References

See Help:References for how to manage references in GONUTS.