Troponin I Encompasses an Extended Troponin C in the Ca2+-Bound Complex: A Small-Angle X-ray and Neutron Scattering Study

Abstract

We have studied the solution structure of skeletal muscle troponin C complexed with troponin I in the presence of calcium using small-angle X-ray and neutron scattering. 4Ca2+.troponin C in the complex has an extended dumbbell shape with a radius of gyration of 23.9 +/- 0.5 A and a maximum linear dimension of approximately 72 A, similar to the values obtained from the crystal structure coordinates of troponin C (Herzberg & James, 1985). Troponin I is even more extended than troponin C with a radius of gyration of 41 +/- 2 A and a maximum linear dimension of approximately 118 A. The centers-of-mass for each component of the complex are approximately coincident (< 10-A separation) as are their long axes, and the troponin I component encompasses the 4Ca2+.troponin C. These data provide new insights into the nature of the conformational arrangement of this important Ca(2+)-sensitive molecular switch.

Invited review: Probing the structures of muscle regulatory proteins using small‐angle solution scattering

“…Upon Ca 21 binding to the regulatory sites of TnC, TnI was observed to undergo a large-scale structural change resulting in a more compact structure with a $ 10% decrease in the radius of gyration R g and a $ 9% increase in the apparent cross-sectional radius of gyration (R c ). By modelling TnI as a two-domain molecule represented by oblate and prolate ellipsoids containing 65% and 35% of the mass, respectively, these researchers concluded there was a decrease in the center-of-mass separation of these domains of $ 15 Å upon Ca 21 addition, a strong indication that TnI acts as a molecular switch consistent with the mechanism proposed by Olah et al 46 Of note, sTnI in the ternary complex 49 was found to be more compact than was observed in for the binary complex, 46,48 suggesting interactions with sTnT were important in determining the structure of sTnI in the complex. Further development of ideas about the proposed Ca 21 -dependent molecular switch in Tn would have to await crystal structure data on Tn (see below).…”

“…46 The sTnC was deuterated, and the SANS contrast variation study showed that 4Ca 21 Á TnC adopts an extended dumbbell shape in the complex with a fully extended interconnecting helix, and that the TnI is even more extended. Analysis of the contrast dependence of the R g values from the complex revealed that the centers of mass of the two components are coincident.…”

Quaternary Structures of a Catalytic Subunit-Regulatory Subunit Dimeric Complex and the Holoenzyme of the cAMP-dependent Protein Kinase by Neutron Contrast Variation

“…Bacterially expressed nonmyristoylated C was not used because of the potential for the myristoyl group to play a role in the structure of the protein that is important for its interactions with R. Bovine heart C␣ was prepared and purified to homogeneity as described previously (26). Deuterated R II ␣ was prepared by expression procedures similar to those used previously for the preparation of other deuterated proteins (27,28). In brief, the murine R II ␣ vector (PET11D), provided by Dr. John Scott (29), was expressed in E. coli BL21DE3 in a mixture of D 2 O and H 2 O containing deuterated algal hydrolysate diluted with nondeuterated amino acids and glucose to give the average levels of deuteration desired (27).…”

Phosphorylase kinase: the complexity of its regulation is reflected in the complexity of its structure

“…As is the case for the delta subunit of Phk, troponin C remains bound to troponin I when the [Ca 2+ ] is lowered to nanomolar levels. Based on small-angle X-ray (SAXS) and neutron (SANS) scattering measurements, a structural model for the troponin C-I complex in solution has been proposed as shown in figure 5 (36,37). In this model troponin C is folded into a rather extended conformation.…”