Functional Characterization of the Secondary Actin Binding Site of Myosin II†

Biochemistry ◽  
1999 ◽  
Vol 38 (46) ◽  
pp. 15078-15085 ◽  
Author(s):  
Juliette Van Dijk ◽  
Marcus Furch ◽  
Chrystel Lafont ◽  
Dietmar J. Manstein ◽  
Patrick Chaussepied
Keyword(s):  
Binding Site ◽  
Myosin Ii ◽  
Functional Characterization ◽  
Actin Binding ◽  
Actin Binding Site

scholarly journals Localization by Segmental Deletion Analysis and Functional Characterization of a Third Actin-binding Site in Domain 5 of Scinderin

1998 ◽  
Vol 273 (6) ◽  
pp. 3661-3668 ◽  
Author(s):  
Monica G. Marcu ◽  
Li Zhang ◽  
Abdelbaset Elzagallaai ◽  
José-Marı́a Trifaró
Keyword(s):  
Binding Site ◽  
Functional Characterization ◽  
Deletion Analysis ◽  
Actin Binding ◽  
Actin Binding Site

scholarly journals Functional characterization of protein 4.1 homolog in amphioxus: Defining a cryptic spectrin-actin-binding site

2013 ◽  
Vol 3 (1) ◽  
Author(s):  
Lixia Wang ◽  
Yuan Wang ◽  
Zhaohe Li ◽  
Zhan Gao ◽  
Shicui Zhang
Keyword(s):  
Binding Site ◽  
Functional Characterization ◽  
Actin Binding ◽  
Protein 4.1 ◽  
Actin Binding Site

scholarly journals Mutational analysis of an actin-binding site of cofilin and characterization of chimeric proteins between cofilin and destrin.

1992 ◽  
Vol 267 (11) ◽  
pp. 7240-7244
Author(s):  
K Moriyama ◽  
N Yonezawa ◽  
H Sakai ◽  
I Yahara ◽  
E Nishida
Keyword(s):  
Binding Site ◽  
Mutational Analysis ◽  
Actin Binding ◽  
Chimeric Proteins ◽  
Actin Binding Site

scholarly journals Domain structure in actin-binding proteins: expression and functional characterization of truncated severin.

1991 ◽  
Vol 112 (4) ◽  
pp. 665-676 ◽  
Author(s):  
L Eichinger ◽  
A A Noegel ◽  
M Schleicher
Keyword(s):  
Amino Acids ◽  
Binding Site ◽  
Binding Sites ◽  
Actin Filaments ◽  
Functional Characterization ◽  
Tryptophan Fluorescence ◽  
Border Region ◽  
Actin Binding ◽  
Capping Proteins ◽  
Actin Binding Site

Severin from Dictyostelium discoideum is a Ca2(+)-activated actin-binding protein that severs actin filaments, nucleates actin assembly, and caps the fast growing ends of actin filaments. Sequence comparison with functionally related proteins, such as gelsolin, villin, or fragmin revealed highly conserved domains which are thought to be of functional significance. To attribute the different activities of the severin molecule to defined regions, progressively truncated severin polypeptides were constructed. The complete cDNA coding for 362 (DS362) amino acids and five 3' deletions coding for 277 (DS277), 177 (DS177), 151 (DS151), 117 (DS117), or 111 (DS111) amino acids were expressed in Escherichia coli. The proteins were purified to homogeneity and then characterized with respect to their effects on the polymerization or depolymerization kinetics of G- or F-actin solutions and their binding to G-actin. Furthermore, the Ca2+ binding of these proteins was investigated with a 45Ca-overlay assay and by monitoring Ca2(+)-dependent changes in tryptophan fluorescence. Bacterially expressed DS362 showed the same Ca2(+)-dependent activities as native severin. DS277, missing the 85 COOH-terminal amino acids of severin, had lost its strict Ca2+ regulation and displayed a Ca2(+)-independent capping activity, but was still Ca2+ dependent in its severing and nucleating activities. DS151 which corresponded to the first domain of gelsolin or villin had completely lost severing and nucleating properties. However, a residual severing activity of approximately 2% was detectable if 26 amino acids more were present at the COOH-terminal end (DS177). This locates similar to gelsolin the second actin-binding site to the border region between the first and second domain. Measuring the fluorescence enhancement of pyrene-labeled G-actin in the presence of DS111 showed that the first actin-binding site was present in the NH2-terminal 111 amino acids. Extension by six or more amino acids stabilized this actin-binding site in such a way that DS117 and even more pronounced DS151 became Ca2(+)-independent capping proteins. In comparison to many reports on gelsolin we draw the following conclusions. Among the three active actin-binding sites in gelsolin the closely neighboured sites one and two share the F-actin fragmenting function, whereas the actin-binding sites two and three, which are located in far distant domains, collaborate for nucleation. In contrast, severin contains two active actin-binding sites which are next to each other and are responsible for the severing as well as the nucleating function. The single actin-binding site near the NH2-terminus is sufficient for capping of actin filaments.

Characterization of an Actin-binding Site within the Talin FERM Domain

2004 ◽  
Vol 343 (3) ◽  
pp. 771-784 ◽  
Author(s):  
Ho-Sup Lee ◽  
Robert M. Bellin ◽  
Diane L. Walker ◽  
Bipin Patel ◽  
Pam Powers ◽  
...  
Keyword(s):  
Binding Site ◽  
Actin Binding ◽  
Ferm Domain ◽  
Actin Binding Site

scholarly journals Characterization of the actin binding site on smooth muscle filamin.

1994 ◽  
Vol 269 (6) ◽  
pp. 4279-4284
Author(s):  
M.C. Lebart ◽  
C. Méjean ◽  
D. Casanova ◽  
E. Audemard ◽  
J. Derancourt ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Binding Site ◽  
Actin Binding ◽  
Actin Binding Site

Characterization of the actin-binding site on the alkali light chain of myosin

1985 ◽  
Vol 830 (3) ◽  
pp. 233-243 ◽  
Author(s):  
Gillian D. Henry ◽  
Monica A. Winstanley ◽  
David C. Dalgarno ◽  
G.Marcus M. Scott ◽  
Barry A. Levine ◽  
...  
Keyword(s):  
Binding Site ◽  
Light Chain ◽  
Actin Binding ◽  
Actin Binding Site

scholarly journals Further characterization of the interaction between the cytoskeletal proteins talin and vinculin

2002 ◽  
Vol 362 (3) ◽  
pp. 761-768 ◽  
Author(s):  
Mark D. BASS ◽  
Bipin PATEL ◽  
Igor G. BARSUKOV ◽  
Ian J. FILLINGHAM ◽  
Robert MASON ◽  
...  
Keyword(s):  
Binding Site ◽  
Intramolecular Interaction ◽  
Binding Activity ◽  
Cytoskeletal Proteins ◽  
Actin Binding ◽  
Yeast Two Hybrid ◽  
Temperature Dependent ◽  
Two Hybrid ◽  
Actin Binding Site

The cytoskeletal protein talin, which is thought to couple integrins to F-actin, contains three binding sites (VBS1—VBS3) for vinculin, a protein implicated in the negative regulation of cell motility and whose activity is modulated by an intramolecular interaction between the vinculin head (Vh) and vinculin tail (Vt) domains. In the present study we show that recombinant talin polypeptides containing the three VBSs (VBS1, residues 498–636; VBS2, residues 727–965; and VBS3, residues 1943–2157) each bind tightly to the same or overlapping sites within vinculin1–258. A short synthetic talin VBS3 peptide (residues 1944–1969) was sufficient to inhibit binding of a 125I-labelled talin VBS3 polypeptide to vinculin1–258, and NMR spectroscopy confirmed that this peptide forms a 1:1 complex in slow exchange with vinculin1–258. Binding of the 125I-labelled VBS3 polypeptide was markedly temperature dependent, but was not inhibited by 1M salt or 10% (v/v) 2-methyl-2-propanol. Attempts to further define the talin-binding site within vinculin1–258 using a gel-blot assay were unsuccessful, but near maximal talin-binding activity was retained by a construct spanning vinculin residues 1–131 in a yeast two-hybrid assay. Interestingly, the talin VBS3 polypeptide was a potent inhibitor of the Vh—Vt interaction, and the VBS3 synthetic peptide was able to expose the actin-binding site in intact vinculin, which is otherwise masked by the Vh—Vt interaction. The results suggest that under certain conditions, talin may be an effective activator of vinculin.

scholarly journals Evidence that a 27-residue sequence is the actin-binding site of ABP-120

1991 ◽  
Vol 266 (20) ◽  
pp. 12989-12993
Author(s):  
A.R. Bresnick ◽  
P.A. Janmey ◽  
J. Condeelis
Keyword(s):  
Binding Site ◽  
Actin Binding ◽  
Actin Binding Site
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