Figure 4

Structural modeling and tryptic digestion of C313Y-MstnPP suggest structural perturbation. A. Orthogonal structural representations of the environment of residue 313 within: (i) The MstnGF crystal structure (green ribbons) highlighting the native C313-C374 disulphide bond (boxed, C313 red; C374 green stick representation) (ii) The modeled C313Y-MstnGF structure with C313 replaced by Tyr showing a superposition of the Tyr sidechain favoured rotamers (Chi 1 angles -60°, 180°, 60°). The intermolecular disulphide (C339-C339') is shown in blue stick representation. The right-hand view is rotated ~90° around the X-axis relative to the left-hand view. B. (i) Limited tryptic proteolysis of C313Y-MstnPP. Reducing (R) and non-reducing (NR) SDS-PAGE showing bands as indicated: M, monomer; D, dimer. Tryptic digestion was performed at 37°C with a myostatin:trypsin ratio of 100:1(w/w) and samples were taken at 0, 0.5, 1, 2, 3, 4 and 18 hour time points. (ii) Quantification of trypsin proteolysis of C313Y-MstnPP and MstnPP [24], as a function of time, normalised to the concentration at time zero.