7.88 Lecture Notes - 7 7.24/7.88J/5.48J The Protein Folding Problem Tropomyosin and S-peptide • Sequence determinants of Coiled Coil Structure • Tropomyosin • Circular Dichroism • Tropomyosin thermal denaturation/renaturation • Chain Recognition, Association, Registration A. Revisit Sequence determination of Coiled Coils H – X – X -H – P – X - P – H – X – X – H – P – X - P- H – X – X - H-P – X -P Lets draw as helical wheel: Note side chain interactions above and below plain: Now lets flip the sequence 18: Sequence: • 1,4 residues: hydrophobic core • 5,7: selectivity, polarity • 2,3, 6: Quaternary interactions; cellular functions Perhaps: But for TM and other coiled coils, easy to see that they encode: Lets review muscle structure: Theta in degrees; Light waves are transverse vibrations; can be vibrating in various planes perpendicular to direction of propagation.
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B. Tropomyosin IN vitro unfolding/refolding of a coiled-coil: Tropomyosin: Properties of Tropomyosin Denaturation/Renaturation of TM Reversibility Coupling of secondary and tertiary structure From thermal denaturation curve: Conformation = f (sequence + environment) Possible pathways Question: Is this conformation/subunit organization encoded by amino acid sequence; perhaps protein folds in association with other components of muscle structure? However, for these structures, only fiber diffraction; no 3-D crystals, no high res structure; not in PDB. In fact in first edition of Branden and Tooze no mention of coiled coils, because not present among globular soluble proteins!! Why tropomyosin: Given that alpha helices are critical conformation of keratin polypeptide chains: What about examining keratin folding into keratin helices using the in vitro methodology of Anfinsen??? Not possible: severe limitations in these systems; Mature fibrils highly covalently crosslinked Solubilization requires severe treatment and does not yield homogenous population of protein chains. Only just recently has it been possible to obtain the precursor forms of keratin chains prior to folding and assembly into hair - from tissue culture and from products of cloned genes: Tropomysoin function and organization: Remind you of structure of striated muscle: • Thin filaments from M band • Thick filaments which are bipolar: Tropomyosin Structure: