Specific sequences determine the stability and cooperativity of folding of the C-terminal half of tropomyosin.


Tropomyosin is a flexible 410 A coiled-coil protein in which the relative stabilities of specific regions may be important for its proper function in the control of muscle contraction. In addition, tropomyosin can be used as a simple model of natural occurrence to understand the inter- and intramolecular interactions that govern the stability of coiled-coils. We have produced eight recombinant tropomyosin fragments (Tm(143-284(5OHW),) Tm(189-284(5OHW)), Tm(189-284), Tm(220-284(5OHW)), Tm(220-284), Tm(143-235), Tm(167-260), and Tm(143-260)) and one synthetic peptide (Ac-Tm(215-235)) to investigate the relative conformational stability of different regions derived from the C-terminal region of the protein, which is known to interact with the troponin complex. Analytical ultracentrifugation experiments show that the fragments that include the last 24 residues of the molecule (Tm(143-284(5OHW)), Tm(189-284(5OHW)), Tm(220-284(5OHW)), Tm(220-284)) are completely dimerized at 10 microm dimer (50 mm phosphate, 100 mm NaCl, 1.0 mm dithiothreitol, and 0.5 mm EDTA, 10 degrees C), whereas fragments that lack the native C terminus (Tm(143-235),Tm(167-260), and Tm(143-260)) are in a monomer-dimer equilibrium under these conditions. The presence of trifluoroethanol resulted in a reduction in the [theta](222)/[theta](208) circular dichroism ratio in all of the fragments and induced stable trimer formation only in those containing residues 261-284. Urea denaturation monitored by circular dichroism and fluorescence revealed that residues 261-284 of tropomyosin are very important for the stability of the C-terminal half of the molecule as a whole. Furthermore, the absence of this region greatly increases the cooperativity of urea-induced unfolding. Temperature and urea denaturation experiments show that Tm(143-235) is less stable than other fragments of the same size. We have identified a number of factors that may contribute to this particular instability, including an interhelix repulsion between g and e' positions of the heptad repeat, a charged residue at the hydrophobic coiled-coil interface, and a greater fraction of beta-branched residues located at d positions. Study holds ProTherm entries: 17649, 17650, 17651, 17652, 17653, 17654, 17655, 17656 Extra Details: 1. 0.5 mM EDTA, 0.5 mM DTT were added in the experiment,2. fragment 143-284 (5 - hydroxytryptophan) coiled-coil protein; intramolecular interactions; C-terminal; interhelix repulsion

Submission Details

ID: jt6V2eB93

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:50 p.m.

Version: 1

Publication Details
Paulucci AA;Hicks L;Machado A;Miranda MT;Kay CM;Farah CS,J. Biol. Chem. (2002) Specific sequences determine the stability and cooperativity of folding of the C-terminal half of tropomyosin. PMID:12167616
Additional Information

Structure view and single mutant data analysis

Study data

No weblogo for data of varying length.
Colors: D E R H K S T N Q A V I L M F Y W C G P

Data Distribution

Studies with similar sequences (approximate matches)

Correlation with other assays (exact sequence matches)

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
100.0 Tropomyosin alpha-1 chain Q5KR49 TPM1_BOVIN
100.0 Tropomyosin alpha-1 chain P84335 TPM1_CHEAU
100.0 Tropomyosin alpha-1 chain P04268 TPM1_CHICK
100.0 Tropomyosin alpha-1 chain P58773 TPM1_COTJA
100.0 Tropomyosin alpha-1 chain P13104 TPM1_DANRE
100.0 Tropomyosin alpha-1 chain P09493 TPM1_HUMAN
100.0 Tropomyosin alpha-1 chain P58771 TPM1_MOUSE
100.0 Tropomyosin alpha-1 chain P42639 TPM1_PIG
100.0 Tropomyosin alpha-1 chain P58772 TPM1_RABIT
100.0 Tropomyosin alpha-1 chain P13105 TPM1_RANTE
100.0 Tropomyosin alpha-1 chain P04692 TPM1_RAT
100.0 Tropomyosin alpha-1 chain Q01173 TPM1_XENLA
100.0 Tropomyosin alpha-1 chain Q5KR47 TPM3_BOVIN
100.0 Tropomyosin alpha-1 chain P06753 TPM3_HUMAN
100.0 Tropomyosin alpha-1 chain A1XQV4 TPM3_PIG