Download Chapter Five * Amino Acids and Proteins

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