Amino Acids: Structure, Properties, and Primary Structure of Proteins - Prof. Catherine C., Study notes of Biology

Download Amino Acids: Structure, Properties, and Primary Structure of Proteins - Prof. Catherine C. and more Study notes Biology in PDF only on Docsity! BCHM 463 Sept 15, 2009 READ Chapter 4. Amino acids Problems 14, 17 General Structure, Fig 4-1. Twenty standard a.a., with codons Table 4-1 explain residue mass, three and one letter symbols. Note pK values for 7 side chains Amino acids are dipolar ions pK values of carboxylic acid ~2.2 (see Table 4-1) pK values of amino groups pK ~9.4 Structures in Table 4-1 are at pH 7.0 pH = pK + log [A-]/[AH] 1 So at physiological pH (~7.4) the amino groups are protonated and the carboxylic acids are in their conjugate base form—carboxylates. Dipolar molecules or zwitterions carry charged groups of opposite sign. Are more soluble in water than in nonpolar solvents B. Peptide bonds incorporate a.a. into polymers. Condensation reaction. H2O is eliminated Draw Fig 4.3 The new amide linkage is also known as a peptide bond. Polymerize to dipeptides, tripeptides, tetra, oligopeptides, polypeptides. Usually just call peptides. When an a.a. is incorporated into a peptide, refer to it as a residue. Residue masses are given in Table 4-1. Linear polymers. One terminal residue has a free amino group. The other a free carboxylic acid group. (see drawing above) By convention, draw amino 2 above discussion. Have to determine experimentally. E. Three letter abbreviations in Table 4-1. Most resemble the full name. Asx means Asp or Asn. Glx means Glu or Gln. In the lab Gln and Asn are easily hydrolyzed to Glu and Asp. Also have a one letter code, that is convenient to use to show sequences of entire proteins. Usually correspond to the first letter of the a.a. name. Show tetrapeptide p. 82 and name chemically and as AYDG Another way to learn some of the twenty: as derivatives of alanine. Draw structures of alanine, Phe, Trp, Ser, Asn, Tyr, Cys, Asp, His 2. Stereochemistry. All are optically active except glycine. The alpha carbons have four different substituents and two potential steric arrangements. HOWEVER, all amino acids have the same sterochemical configuration, the same relative configuration around their alpha atoms. They are classified as L-α amino acids. 5 Figure p.84. Fischer convention : levorotatory config of glyceraldehydes (not R&S convention) Note in Table 4-1 that threonine has a second chiral center—a single defined diastereomer Optically active molecules and stereochemical processes are characteristic of life. Bacteria synthesis D- α amino acids and also variations on the 20 discussed here. These are not products of ribosomal translation, but of secondary metabolism. In the pharmaceutical industry, most drugs have chiral centers. Usually only one enantiomer is active, but the chemical synthesis produces a racemic mixture. fig 4-12 ibuprofen one enantiomer is an anti- inflammatory. The other is inactive. Sell as a mixture fig 4-13 thalidomide one enantiomer is a mild sedative, anti-nasea. The other is a teratogen 3. Amino acid derivatives. a.a side chains are often modified in proteins after the protein is synthesized. Also the N-terminus and C-terminus of proteins can be modified. Often important for function. 6 Fig 4-14 9clicker) salt bridges also stabilize folded proteins Some amino acids and a.a. derivatives have important functions by themselves, independent of proteins . see fig 4-15 for examples. READ: Chapter 5 Proteins: Primary Structure Problems: Note media resources offered by wileyPlus. Kinetic figures, advanced reading Proteins act as catalysts in cell metabolism (enzymes), regulators, communicators, transporters, structural components, etc. 1.polypeptide diversity. We describe a protein’s primary, secondary, tertiary and quaternary structures. Primary structure is the topic today. Primary structure: the a.a. sequence and modifications of the polypeptide chain. Can determine sequences directly or can deduce from gene sequence. Most peptides contain between 100 and 1000 residues. Figure 5-1 insulin. Note disulfide bonds. Is synthesized as a single polypeptide. 7 Limitation is that you have to generate the antibody by exposing an animal to the target protein. So you have to know what your target is. Spectroscopy: The concentration of a substance in solution can be measured by absorbance spectroscopy. Beer-Lambert law. A = log Io/I = εcLcL A is the solution’s absorbance or optical density Io is the intensity of the incident light at a given wavelength λ; I is the transmitted intensity The εcL = absorptivity or extinction coefficient of the solute at λ C is the solute’s concentration The l is the length of the light path in centimeters The value of εcL varies with λ. See Fig 5-4 for absorbance spectra of the aromatic a.a. If the value of εcL is known for the solute/protein then its concentration can be spectroscopically determined. Beckman instrument first commercialization. 10 Proteins do not absorb visible light. Though they may complex a cofactor that is a chromophore. The three aromatic amines absorb well in the UV region and this is used. Not specific to a particular protein of course. But used to assay total protein. Assay at 280 nm. And not as sensitive as needed. Commercial assay kits complex strong dyes with protein to improve spectroscopic sensitivity. Back to purification. Usually by a combination of different procedures that take advantage of the physicochemical properties of the protein of interest to enrich it stepwise. The strategy is to eliminate other components of the mixture. It helps to know something about the target’s solubility, ionic charge, polarity, size and any binding preferences. Table on p. 97 These direct the techniques used. B. salting out separates proteins by their solubility Solubility depends on the polarity of the solvent, the pH, temperature, and on other dissolved salts. High concentrations of salt compete for hydrating water molecules and (ideally) can precipitate the 11 protein. Works best with pH at the pI.Recover by centrifuging. Usually use (NH4)2SO3. Artful. Fig 5-2 pepsin is a gastric enzyme, operates at low pH; histones are basic proteins to bind acidic DNA in chromatin 12