Protein Structure: Understanding Primary, Secondary, Tertiary, and Quaternary Levels, Study notes of Biology

Download Protein Structure: Understanding Primary, Secondary, Tertiary, and Quaternary Levels and more Study notes Biology in PDF only on Docsity! 1 Proteins Dr. Leisha Mullins Sept. 9, 2003 2 Protein Structure • The levels of protein structure - Primary - sequence - Secondary - local structures - Tertiary - overall 3-dimensional shape - Quaternary - subunit organization The Role of the Sequence in Protein Structure All of the information necessary for folding the peptide chain into its "native” structure is contained in the primary amino acid structure of the peptide. What forces determine the structure? • Primary structure - determined by covalent bonds • Secondary, Tertiary, Quaternary structures - all determined by weak forces • Weak forces - H-bonds, ionic interactions, van der Waals interactions, hydrophobic interactions 5 Importance of Primary Sequence Many protein diseases involve a change in the primary amino acid sequence • Sickle Cell Anemia – Hemoglobin: V L S P A D K…….. – Sickle Cell: V L S P A V K……… 6 Secondary Structure • The ordered 3-dimensional arrangements of localized regions of a polypeptide chain • Formation depends only on interactions of the peptide backbone: NH and C=O • The formation of the interactions depends on the allowed rotations around the planer peptide bond 7 Phi and Psi Angles • Two degrees of freedom per residue for the peptide chain • phi (Φ) - Angle about the C(alpha)-N bond • psi (ψ) - Angle about the C(alpha)-C bond • Some values of phi and psi are more likely than others. 10 Stability of alpha helix Amino acid identity and location affects helix stability • Proline disrupts the helix - it creates a bend because of (1) the restricted rotation due to its cyclic structure (2) its α-amino group has no N-H for hydrogen bonding • Several positively or negatively residues adjacent to each other will destablize the helix due to charge – charge repulsion • Adjacent bulky residues will also destabilize the helix 11 β sheets • Polypeptide chains lie adjacent to one another • R groups alternate, first above and then below plane • C=O and N-H groups of each peptide bond are perpendicular to axis of the sheet • C=O---H-N hydrogen bonds are between adjacent sheets and perpendicular to the direction of the sheet • Antiparallel β sheets • Parallel β sheets • Proteins that are primarily β sheets have high content of alanine and glycine 12 Large β sheets • Large β sheets structures can be a mix of parallel and antiparallel β sheets. • Parallel β sheets tend to from large structures > 5 strands • Antiparallel β sheet form smaller structures 15 Supersecondary Structures Specific combinations of α helices and β sheets • βαβ - two parallel strands of β-sheet connected by a stretch of α-helix • αα - two antiparallel α-helices • β meander - an antiparallel sheet formed by a series of tight reverse turns connecting stretches of a polypeptide chain • Greek Key –a repetitive supersecondary structure formed when an antiparallel sheet doubles back on itself 16 Motifs • Repetitive Supersecondary Structures • Motifs give information on the folding of a proteins but NOT about protein function 17 Domains • Very large polypeptide chains tend to fold in two or more clusters known as domains • Small segments usually connect neighboring domains • Domain can be structurally and functionally independent • Domains with similar conformations are often associated with a particular function 20 Fibrous Protein - α Keratin • Found in hair, fingernails, claws, horns and beaks • The basic unit of α keratin is a dimer of α helices • Primary structure of helical rods has a repeating pattern of nonpolar residues that promotes association of helices! • α Keratin forms higher ordered structures – Dimer → Protofilament → Microfibril 21 Curly Hair • Keratin is rich in Cys residues that form disulfide bonds cross-linking the adjacent polypeptide strands • A “permanent” involves reducing and oxidizing the disulfide bonds to change the degree of curl or wave • A “set”, wetting and drying, rearranges the hydrogen bonds between the helices and fibers • So on humid days, a dynamic rearrangement of hydrogen bonds is occurring when hair becomes “frizzy” 22 Fibrous Protein – Silk Fibrion • Found in silk fibers (Silk Fibroin) • antiparallel β-sheets that are aligned parallel to the axis of the fiber • The β-sheets stack to form microcrystalline arrays • Alternating sequence: Gly-X -Gly- X where X is alanine or serine • all glycines on one side and all alanines and serines on other side! 25 Collagen – Triple Helix • The triple helix is much more extended than alpha helix • Every third residue faces the crowded center – only glycine is small enough • Pro permit the sharp twisting of the collagen helix • Fibrils are further strengthened by covalent crosslinks between Lys and His • The amount of crosslinking increases with age 26 Globular Protein • Can contain: - only α helices or only β sheets - significant number of α helices and β sheets • The large number of H bonds in the secondary structural elements act to “neutralize” or “stabilize” the polarity of the peptide backbone • Example – Myoglobin – Functions as an oxygen storage and transport protein in muscles – Has a compact structure • Eight alpha helices – Distribution of amino acids • Nonpolar in interior • Polar exterior • Polar residues found in the interior are part of H bonds 27 Interactions stabilizing Tertiary Structures Native structure is only slightly more stable than the unfolded structure 0.4 kJ /mole/amino acid • 1. Hydrophobic interactions • 2. Electrostatic interactions • 3. Hydrogen bonds • 4. Chemical Crosslinking 30 Denaturation • Non-covalent interactions are weak – Disruptions cause proteins to unfold or denature 31 Renaturation Primary structure dictates the native structure • Under proper conditions denatured proteins can refold to native state • Example – In 1957, Christian Anfinsin showed that ribonuclease A could be denatured and it would spontaneously renature – Ribonuclease A has 4 disulfide bonds in its native state – Ribonuclease A refolds to its native structure even after reduction and reoxidation 32 Protein Folding in vivo Protein Folding conditions in vivo are slightly different than in vitro folding conditions • The redox environment in vivo can be different from in vitro conditions – In cells, sometimes the wrong disulfide bonds are formed – The enzyme protein disulfide isomerase (PDI) is used in cells to reduce and reoxidize disulfide bonds 35 Prion Protein 36 Quaternary Structure of Proteins • Many proteins consist of more than one polypeptide chain • Subunits - different polypeptide chains • The individual subunits associate in a specific geometry for that protein known as the quaternary structure • Subunits interact via non-covalent interactions • Proteins with more than one subunit are called oligomers – Dimer, trimer, tetramer etc. 37 Allosteric Proteins • Subunits interact via non-covalent interactions • Binding of a small molecule, known as an allosteric effector, at a site on one subunit causes changes in the structure of another subunit and therefore the physical characteristic of the overall protein • Hemoglobin is a classic example of an Allosteric Quaternary Protein 40 Heme and Oxygen Binding • O2 bind to the heme group • Polypeptide "cradles" the heme group • CO, NO, H2S can bind to the Fe(II) –heme complex • Their binding affinity is higher than that of O2 – therefore toxic 41 Oxygen Binding to Hemoglobin and Myoglobin • Hemoglobin – 100% saturated in lungs – 50 % saturated in tissues • Myoglobin – 100% saturated in lungs – 100 % saturated in tissues 42 Cooperativity • Oxygen binding changes the Mb and Hb conformation • Total movement of Fe is 1 A • Adjacent subunits' affinity for oxygen increases • This is called positive cooperativity 45 Effect of pH • There are two physiologically consequences to the Bohr effect: 1. In capillaries, high [H+] promote the release of O2 2. In the lungs, oxygenation has the effect of releasing H+ O2 affinity ↑ as pH ↑ or O2 affinity ↓, as pH ↓ 46 Bisphosphoglycerate • BPG, D-2,3-bisphosphoglycerate, is a naturally occurring compound in red blood cells. • Its binds preferentially to the deoxy form • 2,3-BPG bind inside the central cavity of deoxyhemoglobin via ionic interactions • BPG can not bind to the oxy form because the binding pocket is too small 47 Fetal Hemoglobin A fetus receives O2 from its mother’s blood via exchange through the placenta • fetal blood has a higher affinity for O2 than the mother’s Why? • Fetal hemoglobin is different than adult hemoglobin - α2γ2 composition • In the absence of allosteric effectors, the oxygen affinity for HbF is very similar to HbA • HbF has a much lower affinity for BPG than does HbA • Adult HbA has a positively charged His in in the β chain • Fetal HbF has a neutral Ser in the γ chain • This change of a positive charge for a neutral charge reduces the binding affinity of BPG to HbF