Threonine-77 Is a Determinant of the Low-Temperature Conditioning of the Most Abundant Isoform of Tropomyosin in Atlantic Salmon.

The Atlantic salmon survives below 10 °C. The main skeletal muscle is composed of a single isoform of tropomyosin (classified as Tpm1 α-fast) that is >92% identical to the mammalian homologue. How salmon Tpm1 maintains flexibility is investigated by reversing the only full charge substitution; threonine-77(g) in salmon and lysine in other vertebrates. The mutation (Thr-77 to Lys), which falls within a known destabilizing alanine cluster, (i) yields a useful electrophoretic shift in the absence and presence of an anionic detergent, (ii) increases the s of both cooperative transitions (calorimetry, 0.1 M salt, pH 7) [35 °C (minor) and 44 °C (major); Δ = 5 °C, Δ = 3.5 °C], (iii) increases the of CN1A (residues 11-127) to 53 °C (Δ = 13 °C), a value similar to that of mammalian CN1A, (iv) markedly reduces the rate of proteolysis at Leu-169, and (v) weakens the affinity of salmon Tpm1 for troponin-Sepharose. Glu-82(e), the interstrand ionic partner of Lys-77(g), is conserved. The change in ionic interactions at this locus is postulated to be "sensed" in actin period 5 (residues 166-207) and likely beyond. Wild type (acetylated) salmon Tmp1 binds more tightly to F-actin at 4 °C than at 22 °C, which is the opposite of the long-known relationship displayed by the mammalian homologue. All of the evidence indicates that the presence of a neutral 77th amino acid destabilizes a sizable portion of salmon Tpm1 that includes the midregion. Threonine-77 is a key factor in rescuing the thin filament from the peril of cold-induced rigidity.