Isoform-specific interaction of tropomodulin with skeletal muscle and erythrocyte tropomyosins.
Tropomodulin is a tropomyosin-binding protein that localizes to the pointed end of striated muscle thin filaments and caps the pointed end of tropomyosin-actin filaments in vitro. Results from previous studies have suggested that tropomyosin-tropomodulin interactions are isoform-specific. To investigate the molecular basis for the isoform specific interactions of tropomyosin and tropomodulin, we isolated a cDNA for chicken skeletal muscle tropomodulin. The derived amino acid sequence of muscle tropomodulin is 86% identical with that of human erythrocyte tropomodulin, indicating that tropomodulins are highly conserved proteins. Multiple mRNAs seen on Northern blots of chicken skeletal muscle mRNA can be accounted for by multiple polyadenylation signals in the 3'-untranslated region of the cDNA. 125I-Labeled skeletal muscle and erythrocyte tropomyosins were assayed for binding bacterially expressed muscle tropomodulin using a solid phase binding assay. Unexpectedly, skeletal muscle and erythrocyte tropomyosins bound with similar affinities to muscle tropomodulin (Kd values approximately 0.2 microM). However, cross-competition studies using erythrocyte and skeletal muscle tropomyosins indicated that they bound different sites on tropomodulin. Competition studies using recombinant fragments of tropomodulin to map the binding domains for the two tropomyosins demonstrated that residues 6-94 contained the skeletal muscle tropomyosin binding site and residues 90-184 contained the erythrocyte tropomyosin binding site. Binding of different regions of tropomodulin to muscle and non-muscle tropomyosins may permit interaction of tropomodulin with tissue-specific components or may influence the pointed end capping activity of tropomodulin.
Department of Cell Biology, Scripps Research Institute, La Jolla, California 92037.
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Babcock
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Fowler
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001667:Binding, Competitive; D001798:Blood Proteins / Q000378:metabolism; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002645:Chickens; D003001:Cloning, Molecular; D018076:DNA, Complementary; D004912:Erythrocytes / Q000737:chemistry; D015870:Gene Expression; D008840:Microfilament Proteins; D008969:Molecular Sequence Data; D009124:Muscle Proteins / Q000378:metabolism; D011485:Protein Binding; D012333:RNA, Messenger / Q000235:genetics; D011993:Recombinant Fusion Proteins; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D051348:Tropomodulin; D014335:Tropomyosin / Q000378:metabolism*
D001798:Blood Proteins; D002352:Carrier Proteins; D018076:DNA, Complementary; D008840:Microfilament Proteins; D009124:Muscle Proteins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D051348:Tropomodulin; D014335:Tropomyosin