BIOC 460, Spring 2008LEC 4, Peptides/Primary Structure withKey Concepts and Learning Objectives 1LEC 4, Peptides and Primary Structure: Key Concepts• Proteins: primary structure– Peptide bond• amide linkage holding amino acid residues in peptide and proteinpolymers (primary structure of proteins).• Product of condensation of 2 amino acids– Posttranslational modifications of amino acids/proteinsExamples:• hydroxylation of some Pro and Lys residues in collagen (vital forcollagen structure)• carboxylation of some Glu residues (vital for blood clotting)• reversible phosphorylation of some Ser, Thr, and Tyr residues(vital for many regulatory processes)• proteolytic cleavage (vital for some regulatory processes and indigestion of protein nutrients)• disulfide bond formation (vital for structures of some proteins,especially extracellular proteins, and in some coenzyme andenzyme activities)– Sequence of amino acids in protein (primary structure)determines 3-dimensional folding pattern of protein (higherlevels of structure).Key Concepts, continued• Properties of the peptide bond– Partial double bond character of peptide bond -- importantconsequences for 3-dimensional structures of proteins:• planarity of 6-atom peptide unit (peptide bond C=O and N-Hin center, plus αCs on both sides of peptide bond)• no free rotation (cis-trans isomerism)• steric constraints on dihedral angles around backbonebonds for each amino acid residue– N-Cα angle: Φ– Cα-C=O angle: Ψ• Ramachandran diagram: plot of Ψ vs. Φ (angular coordinates) ofamino acid residues in protein(s)Learning Objectives(See also posted Peptide/pH/Ionization practice problems.)• Terminology related to polypeptides: amino acid residue, backbone,side chains, disulfide bonds, conformation, configuration• Write the chemical equation for formation of a peptide bond.• Draw a peptide bond and describe its conformation (3-dimensionalarrangement of atoms).• Explain the relation between the N- and C-terminal residues of apeptide or protein and the numbering of the amino acid residues in thechain, and be able to draw a linear projection structure (like text Fig.2.19) of a short peptide of any given sequence, using the conventionfor writing sequences left to right from amino to carboxy terminus.• Be able to estimate the approximate net charge on a short peptideat any given pH. This requires being given or knowing theapproximate pKa values of the ionizable groups in peptides andproteins (the single α-amino group and single α-carboxyl group on thepeptide, and any ionizable R groups) as well as the chemistry/chargeproperties of those groups in their conjugate acid and conjugate baseforms.Learning Objectives, continued• Explain how the partial double bond character of the peptide bond andsteric effects relate to the conformation of a polypeptide chain, includingwhether peptide bonds in proteins are predominantly cis or trans.• Explain the concept of a 6-atom planar peptide group (from one α-C tothe next α-C), and how one plane can rotate relative to the next plane ina polypeptide backbone, around the Φ and/or Ψ angles.• Explain which bond rotation angle is defined/described as Φ and whichbond rotation angle is described as Ψ. Explain what a Ramachandranplot is, and how it relates to "allowed" combinations of (Φ,Ψ) coordinatesfor proteins.Lecture 4Peptides and Protein PrimaryStructureReading: Berg, Tymoczko & Stryer, 6th ed., Chapter 2, pp. 34-37Practice problems (peptide ionization):http://www.biochem.arizona.edu/classes/bioc460/spring/460web/lectures/PeptidepHPracticeProblems460-08.pdf (also linked in lecturenotes directory)Problems in textbook: chapter 2, pp. 63-64, #6,7,8,13,14Jmol structure showing planarity of peptide bond:http://www.biochem.arizona.edu/classes/bioc462/462a/jmol/peptide/peptide.htmlAnimation of phi (Φ) and psi (Ψ) angles (dihedral angles for an amino acid residue):http://www.biochem.arizona.edu/classes/bioc462/462a/NOTES/Protein_Structure/Rama_animationhtm.htmPROTEINS: PRIMARY STRUCTURE(AMINO ACID SEQUENCE)• Fig. 2.18: Peptide bond formation.condensation of carboxylic acid and amino groupWhat kind of bond/linkage is a peptide bond?(Ester, anhydride, ether, amide, …?)BIOC 460, Spring 2008LEC 4, Peptides/Primary Structure withKey Concepts and Learning Objectives 2• In 55.5 M H2O, will equilibrium lie in the direction ofpeptide bond formation or hydrolysis?• Why aren’t we all just puddles of amino acids?• Thermodynamics (direction) vs. Kinetics (rate)Terminology: N-terminus, C-terminus, A.A. residueNumberingBackbonePolaritySequenceCompositionBackbone of peptide• Hydrogen bonding potential• Nomenclature:L-aspartyl-L-phenylalanine methyl ester (Asp-Phe-O-CH3) -- common names?• Oligo- vs. Polypeptide• Mass in daltons (amu) or kilodaltons (kD)• Mean residue weight ~110 daltonsPosttranslational modification of proteins• Chemical modification after protein synthesis• Modification carried out by specific enzymes• Examples:– Hydroxylation of Pro or Lys• 4-hydroxyproline(Hyp)• Enzyme: prolyl hydroxylase• Collagen/connective tissue• Vitamin C required for the hydroxylase• What disease results from vitamin C deficiency?Posttranslational modification of proteins• Carboxylation of specific Glu residues on gamma(γ) carbon• Enzyme: glutamate carboxylase• γ-carboxyGlu (gla)• Required for function of several blood clotting enzymes• Involved in Ca2+ binding/ co-localization with platelets at wound sites• Vitamin K required for recycling active form of carboxylase) What would be the effect of a deficiency in Vitamin K?Posttranslational modification of proteins• Phosphorylation of specific Ser (S), Thr (T), or Tyr (Y)residues by (enzymes): protein kinases (add PO32–)Phospho-SerPhospho-Tyr• Modification removed by (different enzymes): protein phosphatases (remove PO32–)What type of bond links the phosphate to the Ser or Thr or Tyr side chain?BIOC 460, Spring 2008LEC 4, Peptides/Primary Structure withKey Concepts and Learning Objectives 3Disulfide Bond Formation(oxidation of cysteinyl residues)• --> covalent crosslinks between Cys residues• Oxidation (loss of 2 e–) makes diS bond.• Reduction (gain of 2 e–) breaks diS bond. Berg et al., Fig. 2.21Amino acid sequence (primary structure)• Sequence (order) of amino acids in chain• Product of translation on a ribosome
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