Polymorphism in tropomyosin structure and function

Abstract

Tropomyosins (Tm) in humans are expressed from four distinct genes and by alternate splicing >40 different Tm polypeptide chains can be made. The functional Tm unit is a dimer of two parallel polypeptide chains and these can be assembled from identical (homodimer) or different (heterodimer) polypeptide chains provided both chains are of the same length. Since most cells express multiple isoforms of Tm, the number of different homo and heterodimers that can be assembled becomes very large. We review the mechanism of dimer assembly and how preferential assembly of some heterodimers is driven by thermodynamic stability. We examine how in vitro studies can reveal functional differences between Tm homo and heterodimers (stability, actin affinity, flexibility) and the implication for how there could be selection of Tm isomers in the assembly on to an actin filament. The role of Tm heterodimers becomes more complex when mutations in Tm are considered, such as those associated with cardiomyopathies, since mutations can appear in only one of the chains.

Fig. 1

Sequence alignment of the putative trigger motifs from exon7 and 8 of the four human TPM genes. Lower case letters indicate heptad position; residues matching consensus sequence are in bold; x, any residue; c, charged residue; h, hydrophobic residue, residue in bold square indicates the position of recently discovered dilated cardiomyopathy (DCM) mutation Asp230Asn in skα-Tm

Fig. 2

a Helix unfolding of gizzard α-Tm (GG), rabbit skeletal α-Tm (RR) and an equal mixture of GG+RR. Note that: (1) GG is less stable than RR; (2) in the mixture, GG starts to unfold first, but at 39 °C refolds with a different profile from GG or RR; (3) on slow cooling and reheating (left right arrow) this intermediate stability is maintained. b Helix unfolding of dimers of rabbit skeletal α-Tm (RR) and chicken gizzard α-Tm (GG) and the product (RG) after the heating/cooling cycle of RR+GG. RG is not an equal mixture of RR and GG. c Gels of DTNB-cross-linked samples of rabbit skeletal α-Tm (R) and chicken gizzard α-Tm (G). RR+GG mixtures were treated at room temperature with 1 mM DTNB for several hours before and after thermal unfolding/refolding (see CD run). Conditions: 0.03 mg/ml Tm in 0.5 M NaCl, 5 mM Hepes buffer pH 7.3

Fig. 3

Thermal unfolding of Tm homo and heterodimers. Normalised unfolding profiles of 7 μM cross-linked homo and heterodimers of sk-Tm. a skαα-Tm (black line), skββ-Tm (dashed lines) and skαβ-Tm (grey line); data from Kalyva et al. (2012). b skαα-wt Tm (black line) and the homo (grey line) and heterodimer (dashed line) of the E180G cardiomyopathy mutation. Data from Janco et al. 2012. Each plot is the average of three repeated melting curves on the same sample. Buffer conditions: 0.5 M KCl, 5 mM MgCl₂, 20 mM potassium phosphate buffer, pH 7.0

Fig. 4

End view of a Tm dimer binding to a partner protein such as Tn. The binding site is asymmetric. If the two Tm chains are identical there may be no preference for how Tn binds to the two Tm chains but if the two chains differ in a heterodimer then the two binding modes can differ