Molecular cloning and functional characterization of mouse coactosin-like protein.
Coactosin was first isolated from Dictyostelium discoideum and, as reported, human coactosin-like protein (CLP) was identified in a yeast two-hybrid screen using 5-lipoxygenase (5LO) as a bait. A mouse CLP (mCLP) cDNA clone was identified among EMBL/GenBank EST sequences. The derived amino acid sequence (142 residues) was 95.1% identical with human CLP. Here, we also show that mCLP interacts with actin and 5LO in the two-hybrid system. High-speed cosedimentation assays and GST-binding assays confirmed these protein interactions. In chemical cross-linking experiments, one molecule of mCLP was covalently linked to either one subunit of actin or one molecule of 5LO. The mCLP-F-actin and mCLP-5LO associations were pH-insensitive and Ca(2+)-independent. However, association with actin was best observed at low salt concentrations, while association with 5LO was favored by salt, indicating different binding characteristics.
Doucet
Johanne
J
Department of Medical Biochemistry and Biophysics, Karolinska Institute, Stockholm, S-171 77, Sweden.
Provost
Patrick
P
Samuelsson
Bengt
B
Rådmark
Olof
O
D000199:Actins / Q000235:genetics / Q000378:metabolism; D000818:Animals; D001094:Arachidonate 5-Lipoxygenase / Q000235:genetics / Q000378:metabolism; D002118:Calcium / Q000737:chemistry / Q000494:pharmacology; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D005982:Glutathione Transferase / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D015151:Immunoblotting; D051379:Mice; D008840:Microfilament Proteins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D011189:Potassium Chloride / Q000737:chemistry / Q000494:pharmacology; D011485:Protein Binding / Q000187:drug effects / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D020798:Two-Hybrid System Techniques
D000199:Actins; C485551:Cotl1 protein, mouse; D018076:DNA, Complementary; D008840:Microfilament Proteins; D011993:Recombinant Fusion Proteins; D011189:Potassium Chloride; D001094:Arachidonate 5-Lipoxygenase; D005982:Glutathione Transferase; D002118:Calcium