Biol 2012 Exploring Proteins PDF

Biol 2012 Exploring Proteins Jörn Werner 1 Today’s Lecture Protein Architecture: 1. Primary, Secondary, tertiary and quaternary structure 2. Ramachandran plot 3. Secondary structural elements 4. General features of tertiary structure in soluble proteins 5. Posttransla...

Biol 2012 Exploring Proteins Jörn Werner 1 Today’s Lecture Protein Architecture: 1. Primary, Secondary, tertiary and quaternary structure 2. Ramachandran plot 3. Secondary structural elements 4. General features of tertiary structure in soluble proteins 5. Posttranslational Modifications  Hierarchical view of protein structure Sanger X-ray diffraction X-ray crystallography Gel-filtration, sequencing patterns Scattering, 3 X-ray crystallography Part 1: Primary Structure 4 The Alphabet 20 amino acids are the building blocks of proteins Diverse functional groups every amino acid is special may be chemically modified classified according to chemical properties Non-polar 5 Three facts about the peptide bond A) Condensation reaction B) Partial double bond character C) Planarity (w=180) w 6  Peptide Bond 2: Trans configuration is preferred (energetically much lower) for all amino acids Special Property 1: Proline Unfolded peptides: 2/3 trans & 1/3 cis Folded proteins: in some cases peptide bonds are in cis. they are target of the enzyme: proline cis/trans isomerase 7  Peptide backbone conformation w Three dihedral angles determine the backbone conformation of a protein 1) w (peptide bond) close to 180 2) f (pronounce phi) rotation around the NC bond 3) y (pronounce psi) rotation around the CC’ bond 8  Ramachandran plot of protein backbone conformation y f Steric clashes between next neighbour residues greatly constrain backbone conformation 9  Regular secondary structure can be read of the Ramachandran plot Right-handed a-helical backbone conformation, Extended backbone conformation, b-strand 10  Multiple b-strands can associate via backbone H- bonds Two b-strands in an antiparallel arrangement Two b-strands in a parallel arrangement 11 Parallel and antiparallel b-strands can be mixed leading to various b-sheet configurations 12 The direction of the polypetide chain can be reversed by turns and loops Tight turns: Loops: various types with defined structure More than 5 amino acids Typically, less than 5 amino acids Less regular structural elements 13  Example of a mainly b-sheet protein Fatty acid binding protein PDB: 1FDQ LIGAND: DOCOSA-4,7,10,13,16,19-HEXAENOIC ACID 14 a-helices have a conformation with a distinct backbone H- bond pattern 15 a-helices: variation on a theme f,y Turn per # aa Translation H-bond pattern amino per along helix axis acid turn per turn (pitch) regular -60°,-45° 100 ° 3.6 5.4 Å CO(i), NH(i+4) a-helix 3-10 −49°,−26° 120° 3.0 6.0 Å CO(i), NH(i+3) helix p helix -55°,-70° 87° 4.1 4.7 Å CO(i), NH(i+5) 16 Periodic structure of helices 1: Amphipathic Helices hydrophobic hydrophilic 17 Amphipathic helices in water and lipid bilayers water lipid bilayer Hydrophobi Hydrophili c side out c side out 18 Periodic structure of helices 2: Leucine zippers The structure of the AP-1/DNA complex. AP-1 is a dimer formed by Jun and its homologous protein Fos. It contains a leucine zipper motif (blue color) where two a helices look like a zipper with leucine residues (red color) lining on the inside of the zipper. 19 Heptad repeats and coiled-coiled structures Depending on the identity of amino acids in position 4 and 7 20  Some globular proteins predominantly contain one type of secondary structure 21 Protein tertiary structure is stabilized by multiple weak interactions Protein folding (structure formation) is driven by the hydrophobic effect Entropy driven effect: Folding occurs when of gain of entropy upon folding of solute (water) overcomes the loss of conformational entropy of protein. Tertiary structure stabilized by Van der Waals interactions Ionic interactions H-bonds 22  Myoglobin (Oxygen Storage) water soluble protein Ribbon diagram Space filling model Exterior of protein Interior of protein blue: charged residues yellow: hydrophobic residues 23 Some protein structures are stabilized by disulfide bonds Special Property 2:Cysteine/Cystine 24  Quaternary structure: Polypeptide chains can assemble into multi-subunit structures Homo dimer: DNA binding protein Cro of bacteriophage a2b2 tetramer hemoglobin l 25  Protein post-translational modifications A number of different amino acid side chains can be covalently modified to alter protein function - Typically, reversible - enzymatically catalysed - enhance functional diversity - local, temporal control of function 26  Example: Phosphorylation of OH groups Special amino acids (3): Ser, Thr, Tyr Reaction is naturally very slow. Hence, catalysed by kinases and removal by phosphatases 27 Catalysis of phosphorylation is mediated by a kinase domain N-lobe mainly B-sheet g-phosphate from ATP transferred to Tyr, Ser, Thr ATP biding cleft (a) of kinase itself (cis): auto-phosphroylationr C-lobe (b) target protein (trans): mainly a-helical cross-phosporylation  Autophosphorylation involves structural change Inactive Active low rate of high rate of phosphorylation phosphorylation Activation Loop Activation Loop 29 Example: Insulin Receptor Kinases (IRK)  Phosphorylation controls cell signalling A cascade of protein-serine/threonine kinases link G-protein-coupled receptors (GPCR) to the control of glycogenolysis Krebs,E.G. Fisher, E.H Nobel lecture 1992 https://www.nobelprize.org/prizes/ medicine/1992/krebs/lecture/ Pawson 2005, TIBS 30,286 Summary Hierarchical view of protein structure is a result of the techniques employed to study them Functional diversity of amino acids provide rich functional repertoire Every amino acid has special powers Post translational modifications enhance repertoire and enable “fast” “temporal and spatial” control of protein function backbone conformation is restricted by local steric clashes enhances formation of folded structures a-helices and b-pleated sheets have regular H-bond patterns Amino acid sequence motifs may specify specific tertiary structures