Peptides and Proteins - Biochemistry - Lecture Slides, Slides of Biochemistry

Download Peptides and Proteins - Biochemistry - Lecture Slides and more Slides Biochemistry in PDF only on Docsity! Biomolecules: Peptides and Proteins Lecture 5, Medical Biochemistry Docsity.com Lecture 5 Outline • Overview of amino acids, peptides and the peptide bond • Discuss the levels of protein structure • Describe techniques used for analysis of proteins Docsity.com Protein Structure Levels • PRIMARY: the linear sequence of amino acids linked together by peptide bonds • SECONDARY: regions within polypeptide chains with regular, recurring, localized structure stabilized by H-bonding between constituent amino acid residues Docsity.com Protein Structure Levels (cont) • TERTIARY: the overall three- dimensional conformation of a protein • QUATERNARY: the three-dimensional conformation of a protein composed of multiple polypeptide subunits • THE PRIMARY AMINO ACID SEQUENCE IS THE ULTIMATE DETERMINANT OF FINAL PROTEIN STRUCTURE Docsity.com Ex: INSULIN Disulfide bonds Form between two intra- or interchain cysteine residues, product called cystine - Stabilizes/creates protein conformation - Prevalent in extracellular/ secreted proteins Docsity.com 2o Structure: a-helix each oxygen of a carbonyl group of a peptide bond forms a H-bond with the hydrogen atom attached to a nitrogen in a peptide bond 4 amino acids further along the chain; very stable structurally; prolines will disrupt helix formation Docsity.com End-on view of a-helix Docsity.com Parallel Anti-Parallel b-sheet In this secondary structure, each amino acid residue is rotated 180o relative to its adjacent residue. Occur most commonly in anti-parallel directions, but can also be found in parallel. H-bonds between adjacent chains aid in stabilizing the conformation. Docsity.com Domains, examples: Saddle b-Barrel Bundle Docsity.com Ex: Tertiary Structure Ex: Quaternary Structure Docsity.com Myoglobin b-subunit Hemoglobin Docsity.com Myoglobin Properties • At the tertiary level, surface residues prevent one myoglobin from binding complementarily with another myoglobin; thus it only exists as a monomer. • Each monomer contains a heme prosthetic group: a protoporphryin IX derivative with a bound Fe2+ atom. • Can only bind one oxygen (O2) per monomer • The normal physiological [O2] at the muscle is high enough to saturate O2 binding of myoglobin. Docsity.com Heme Structure CoO \ A reine Protein-Heme Complex with bound oxygen Hemoglobin Properties • At the tertiary level, the surface residues of the a and b subunits form complementary sites that promote tetramer formation (a2b2), the normal physiological form of hemoglobin. • Contains 4 heme groups, so up to 4 O2 can be bound • Its physiological role is as a carrier/transporter of oxygen from the lungs to the rest of the body, therefore its oxygen binding affinity is much lower than that of myoglobin. • If the Fe2+ becomes oxidized to Fe3+ by chemicals or oxidants, oxygen can no longer bind, called Methemoglobin Docsity.com SDS-Polyacrylamide Gel (cont) Separation of proteins based on their size is linear in relation to the distance migrated in the gel. Using protein standards of known mass and staining of the separated proteins with dye, the mass of the proteins in the sample can be determined. This is useful for purification and diagnostic purposes. Docsity.com Gel filtration Separation is based on protein size. Dextran or polyacrylamide beads of uniform diameter are manufactured with different pore sizes. Depending on the sizes of the proteins to be separated, they will enter the pore if small enough, or be excluded if they are too large. Hydrophobic Chromatography Proteins are separated based on their net content of hydrophobic amino acids. A hydrocarbon chain of 4-16 carbons is the usual type of resin. Docsity.com Separation of proteins based on the net charge of their constituent amino acids. Different salt concentrations can be used to elute the bound proteins into tubes in a fraction collector. As shown below, resins for binding (+) or (-) charged proteins can be used Ion Exchange Chromatography Docsity.com