THE SEQUENCE OF THE ALTERNATIVELY SPLICED 6TH EXON OF ALPHA-TROPOMYOSIN IS CRITICAL FOR COOPERATIVE ACTIN-BINDING BUT NOT FOR INTERACTION WITH TROPONIN

Tropomyosins, a family of highly conserved coiled-coil actin binding p roteins, can differ as a consequence of alternative expression of seve ral exons (Lees-Miller, J., and Helfman, D. (1991) BioEssays 13, 429-4 57). Exon 6, which encodes residues 189-213 in long, 284-residue tropo myosins, has two alternative forms, exon 6a or 6b, both highly conserv ed throughout evolution, In alpha-tropomyosin, exon 6a or 6b is not sp ecific to any one of the nine isoforms, Exon 6b encodes part of a puta tive Ca2+-sensitive troponin binding site in striated muscle tropomyos ins, suggesting that the exon 6-encoded region may be specialized for certain tropomyosin functions, A series of recombinant, unacetylated , tropomyosin exon 6 deletion and substitution mutants and chimeras was expressed in Escherichia coli to determine the requirements of exon 6 for tropomyosin function, Functional properties of the tropomyosins w ere defined by actin affinity measured by cosedimentation, troponin T affinity using a newly developed biosensor assay, and regulation of th e actomyosin MgATPase. The region of tropomyosin encoded by exon 6 aff ects actin affinity but not thin filament assembly, troponin T binding , or regulation with troponin, The tropomyosins with exon 6a or 6b fun ction normally whether a striated muscle exon 9a or smooth/non-muscle exon 9d is present, However, the effect of deleting 21 amino acids enc oded by exon 6 or replacing it with a GCN4 leucine zipper sequence dep ends on the COOH-terminal sequence.