Protein Structure and Function Lecture 3 Notes: Protein Folding Part 3 Post-Translational and Co-Translational Folding - Kint is the initiation of translation rate. KF and Ku are the bulk folding and unfolding rates. KFi and Kui are the folding and unfolding rates following the release of the ith amino acid. - Translation may 'stall' at specific mRNA positions, which will improve folding. Translation may be halted after SP emergence from the ribosome, until it binds SRP receptor at the ER membrane and docks the nascent chain on the Sec61 translocon. - Co-translational folding within the ribosome is exemplified in the zinc-finger domain, deep inside the ribosomal exit tunnel. - Signal peptides target newly synthesised proteins to the ER membrane for the secretory pathway - The intracellular environment is very different than in vitro folding experiments due to high concentrations of macromolecules. This presents challenges to protein folding including: (a) Macromolecular crowding inside the cell (volume exclusion) (b) Hindered mobility and sticky neighbours (c) Macromolecular crowding affects viscosity (d) Macromolecular crowding favours compaction therefore shifts denatured and intermediate ensembles towards less extended states (e) Other proteins and RNA may affect folding through physical interaction (f) Interaction with membranes and other organelles (g) Molecular chaperones required to assist folding - Many diseases are the result of defective (i.e. misfolded) proteins: (a) Alzheimer's disease (amyloid-ß-protein) (b) Amyotrophic lateral sclerosis (superoxide dismutase) (c) Huntington's disease (Huntingtin with polyglutamate expansion) (d) Lysozyme amyloidosis (lysozyme, forms fibrils) (e) Hereditary renal amyloidosis (fibrinogen) (f) Parkinson's disease (a-synuclein) (g) Transmissible spongiform encephalopathiesffTSEs (prion protein) Protein Geometry - Conformational flexibility is limited to :O: :O :0-0 0 .. rotations along the two bonds of Ca atoms. C. R' R R- N- R- - Peptide units do not involve different side chains. There is a double bond character in :Z-I N + H 1 Kint KA2 i-1 0 KAI KU. K? 1 KUI-, |K?1-1 5 KAH1 R1 H 0 R3 H 0 I 4 N N 0 N O H- 1 H R2 R4 Kui | KEI Kui Kint KA.2 2 KAI KAI+1 planar peptide bond segments KA.42
amide (peptide) bonds between carbon (central carbon) and nitrogen (from amide group) which is very rigid. Presence of nitrogen electron pair delocalisation into the carbonyl group. - Conformation of the whole main chain is completely determined when ¢ (phi) and ? (psi) angles are defined. The only degrees of freedom are rotations around the Ca covalent bonds, so these angles are present at N=C' ( bond angle) and Ca-C' (y bond angle). - Ramachandran plots indicate allowed conformations of polypeptides. ¢ and y angles are plotted against each other using observed values for all amino acids (except glycine) from known X-ray structures. - Glycine adopts a wider range of conformations as it allows unique combinations of ¢ and y angles. Glycine has a special role in folding as it allows unusual conformations and flexibility. - Hovmöller et al. plotted over 200,000 amino acids in over 1000 protein subunits into Ramachandran plots. All