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Download Chapter Five * Amino Acids and Proteins
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5.1 Identity and Roles of Amino Acids • Contain an amine group – Primary amine group • Contain carboxyl group – Referred to as the alpha carbon • R group – Variable group – 20 different groups 5.1 Identity and Roles of Amino Acids • Stereochemistry – All amino acids but one are chiral – Stereochemistry is L 5.2 Amino Acid Individuality: The R Groups • Polarity differs – Polar Amino Acids • Neutral • Acidic • Basic – Nonpolar • • • • Alkyl Branched Aromatics Unique 5.2 Amino Acid Individualities • Functional Groups – Acid Groups • Aspartate, Glutamate, Cysteine, Histidine, Tryosine – Basic Groups • Lysine, Arginine, Histidine – Hydroxyl Groups • Serine, Threonine, Tyrosine – Sulfur Groups • Cysteine, Methionine 5.3 Acid – Base Properties and Charge • Titration and Net Charge – Henderson – Hasselbach equation illustrates the relationship between acid – base dissociation and charge – Amino acids have more than one dissociable group 5.3 Acid – Base Properties and Charge • Zwitterions – Two charged groups, but overall charge is zero – Most amino acids are in this form • Zwitterions lead to altered pK values 5.3 Acid – Base Properties and Charge • Isoelectric point – pH at which zwitterion is exactly zero – Referred to as pI • R groups can have dissociable groups – Acid, basic and sulfhydro groups 5.4 The Peptide Bond • Covalent bond between amino group and alpha carboxyl group • Dehydration reaction • Referred to as amide or peptide bond 5.4 The Peptide Bond • Resonance properties – O, C and N bonds are delocalized – Creates partial double bond 5.4 The Peptide bond • Amine and carboxyl groups are no longer acids or bases • N- terminal end is only amino group not in peptide bond • C- terminal end is only carboxyl group not in peptide bond 5.5 Peptides and Proteins • Peptide – short chain or less than 25 amino acids – Can have biological properties • Protein – more than 25 amino acids • Referred to as macromolecules – Are polymers 5.6 Levels of protein structure • 4 levels of hierarchy – Primary – Secondary – Tertiary – Quaternary 5.6 Levels of Protein Structure • Primary – Sequence of amino acids – Complete description for peptides – Determines its complete spatial arrangement 5.6 Levels of Protein Structure • Secondary – Regular repeating structures – Three main types • Alpha helix – spiral chain – – – – Hydrogen bonds satisified intramolecular R groups determine water solubility Certain amino acids favor Certain amino acids are helix breakers 5.6 Levels of Protein Structure – Three main types cont. • Beta sheets – Resembles corrugate plate – Hydrogen bonds between backbone peptide bonds – Anti parallel and parallel forms • Loops and Random Coils – Indeterminate in structure – Serve to link other secondary structures 5.6 Levels of Protein Structure • Tertiary – Three dimensional arrangement • Quaternary – Three dimensional arrangement of multi subunit proteins – Individual protein chains 5.6 Levels of Protein Structure • Domains – Combination of secondary structural elements – Also referred to as motifs or super secondary structure 5.6 Levels of Protein Structure • Domain examples – TIM domain – SH2 domain – PTB domain – NAD binding domain – EF hand domain 5.7 – Protein folding • Two studies helped increase our understanding • RNase experiment – Enzyme was inactivated with heat and then cooled – On cooling activity resumed – Suggest we can denature and refold a protein • GroEL experiment – Helps protein fold correctly – Increases rate – Often referred to as a chaperone protein 5.8 Oxygen binding in myoglobin and hemeglobin • Myoglobin (Mb) and Hemeglobin (Hb) are oxygen binding proteins in mammals • Similar protein structures – Mb – single chain • Tightly bound to heme – Hb – four chains • 2 Alpha and 2 Beta chains • 4 heme molecules bind 5.8 Oxygen binding in myoglobin and hemeglobin • Heme – Prosthetic group that binds oxygen – Four linked pyrrole rings – Each Nitrogen is chelated to central Iron 5.8 Oxygen binding in myoglobin and hemeglobin • Binding of oxygen occurs differently – Fractional saturation of oxygen – Y = [MbO2]/([Mb] + [MbO2]) – Mb stores oxygen in muscle cells • Releases when levels become depleted 5.8 Oxygen binding in myoglobin and hemeglobin – Hb binds oxygen with positive cooperatively • When one oxygen binds, it helps the other oxygen bind • S shaped curve • Saturated with oxygen in the lungs and releases in tissue capillaries 5.8 Oxygen binding in myoglobin and hemeglobin • Bohr effect – Diminished binding of oxygen when increased pH – Adaptation at high altitudes • 2,3 Bisphophoglycerate – Lowers binding of O2 to hemeglobin – Provides more oxygen to tissues 5.9 Protein purification and Analysis • Purification – Extraction comes first • Properties must be intact – Must have way to measure protein – Must minimize denaturation 5.9 Protein purification and Analysis – Physical methods are often used – Fractionation depends on differences between proteins • Solubility • Size • Charge 5.9 Protein purification and Analysis • Types of chromatography that may be used – Ion exchange column chromatography – Size Exclusion chromatography – High Pressure Liquid Chromatography – Gas Liquid Chromatography – Affinity Chromatography 5.9 Protein purification and Analysis • Analysis – UV light • Tryptophan and tyrosine absorb • Can estimate amount of protein – NMR • Great for small molecule 5.9 Protein purification and Analysis • Analysis cont. – X-Ray • Produce pattern of diffraction • Mathematical manipulation – Mass Spectrometry – Electrophoresis • Can identify various proteins present • Can be a test of purity