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PDBsum entry 1kq5
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Signaling protein
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PDB id
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1kq5
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Contents |
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* Residue conservation analysis
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References listed in PDB file
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Key reference
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Title
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Mutant structures of the actin-Binding domain of cyclase associated protein (cap) from saccharomyces cerevisiae
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Authors
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A.A.Fedorov,
T.Dodatko,
D.A.Roswarski,
S.C.Almo.
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Ref.
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To be Published ...
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Secondary reference #1
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Title
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Two separate functions are encoded by the carboxyl-Terminal domains of the yeast cyclase-Associated protein and its mammalian homologs. Dimerization and actin binding.
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Authors
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A.Zelicof,
V.Protopopov,
D.David,
X.Y.Lin,
V.Lustgarten,
J.E.Gerst.
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Ref.
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J Biol Chem, 1996,
271,
18243-18252.
[DOI no: ]
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PubMed id
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Figure 4.
Fig. 4. The carboxyl terminus of both CAP and MCH1 binds
actin. A, full-length MCH1 and CAP bind actin. Lysates from  cap cells
(SKN50) expressing either Myc-tagged CAP or Myc-tagged MCH1 were
subjected to immunoprecipitation with anti-Myc antibody (9E10).
Specific protein interactions were blocked by the addition of
excess Myc peptide (50 µg). Immune complexes were resolved
on SDS-acylamide gels and transferred to nylon membranes.
Immunoblots were probed with an anti-actin monoclonal antibody
(20 µg/ml) (Boehringer Mannheim). B, the carboxyl-terminal
domain of CAP and MCH1 binds actin. Lysates from cap cells
expressing various Myc-tagged domains of CAP, or the
carboxyl-terminal of MCH1, were subjected to
immunoprecipitation. Immunoblots were incubated with an
anti-actin monoclonal antibody (20 µg/ml). Antigen
detection was performed by ECL chemiluminescent assay.
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Figure 5.
Fig. 5. The actin binding and dimerization functions of CAP
are mediated by separate domains. A, the last 27 amino acids of
CAP are essential for actin binding. cap yeast
cells (SKN32) bearing plasmids expressing HA-CAP (YCpADH-HACAP)
and Myc-CAP1-498 (pADH-mycCAP 11) were
grown to log phase, prior to harvesting, and lysate preparation.
B, CAP1-498 forms homodimers. cap yeast
cells (SKN32) bearing plasmids expressing HA-CAP1-498
(pTADH-HACAP 11) and
Myc-CAP1-498 (pADH-mycCAP 11) were
grown to log phase, prior to harvesting and lysate preparation.
Immunoprecipitations were carried out, as described under
``Experimental Procedures,'' using either anti-HA ascites
(12CA5) or anti-Myc antisera (9E10). Specific protein-protein
interactions were blocked by the addition of either excess HA
(30 µg) or Myc peptide (30 µg) to
immunoprecipitation reactions (+peptide). Detection of tagged
CAP molecules or actin was accomplished using the appropriate
monoclonal antibodies (anti-actin (6 µg/ml), anti-HA
ascites fluid (1:5000), and anti-Myc (1:5000)). Lanes marked TCL
contained 75 µg of total cell lysate. Antigen detection
was performed by ECL chemiluminescent assay.
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The above figures are
reproduced from the cited reference
with permission from the ASBMB
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