horton 3-wa
TRANSCRIPT
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Principles of BiochemistryFourth Edition
Chapter 3Amino Acids and the Primary
Structures of Proteins
Copyright © 2006 Pearson Prentice Hall, Inc.
Horton • Moran • Scrimgeour • Perry • Rawn
Dr. Wisam Alsamah
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Biological Functions of Proteins
1. As enzymes (catalysts)2. Storage and transport (hemoglobin)3. Provide support and shape (collagen)4. Mechanical work (muscles contraction)5. Decoding information (translation)6. As hormones 7. Specialized functions (antibodies)
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General Structure of Amino Acids
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General Structure of Amino Acids
Zwitterions
Chiral OrAsymmetric
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General Structure of Amino Acids
Ball-and-stick model ofserine
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General Structure of Amino AcidsStereoisomers (enantiomers)
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General Structure of Amino Acids
The RS system
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General Structure of Amino Acids
The RS system
Clockwise R configuration
Counterclockwise S configuration
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Structures of the 20 common Amino Acids
A. Aliphatic R Groups
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A. Aliphatic R Groups
Structures of the 20 common Amino Acids
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A. Aliphatic R Groups
Structures of the 20 common Amino Acids
Pyrrolidine ring
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A. Aliphatic R Groups
Structures of the 20 common Amino Acids
●Valine, Leucine, and isoleucine are known as branched-chain amino acids
●All three amino acids are highly hydrophobic
●Pyrrolidine ring of proline restricts the geometry of polypeptides
●Proline is much less hydrophobic than the three amino acids
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B. Aromatic R Groups
Structures of the 20 common Amino Acids
(Benzyl) (Phenol) (Indol)(260 nm) (280 nm) (280 nm)
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C. Sulfur-Containing R Groups
Structures of the 20 common Amino Acids
Disulfide bond
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C. Sulfur-Containing R Groups
Structures of the 20 common Amino Acids
Nonpolar Methyl thioether
Sulfhydryl
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D. Side Chains with Alcohol Groups
Structures of the 20 common Amino Acids
α- carbon
β- carbon
Stereoisomers
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E. Basic R Groups
Structures of the 20 common Amino Acids
Imidazole ring
Guanidinium ion
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F. Acidic R Groups and Their Amide Derivatives
Structures of the 20 common Amino Acids
Amide
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F. Acidic R Groups and Their Amide Derivatives
Structures of the 20 common Amino Acids
●Aspartate, and glutamate are dicarboxylic amino acidsand have negatively charged hydrophilic side chains at pH 7.
●These amino acids are often found on the surface of proteins.
●Monosodium glutamate (MSG) is used as a flavor enhancer.
●The polar amide groups of these amino acids can form hydrogen bonds with atoms in the side chain of other polar amino acids.
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G. The Hydrophobicity of Amino Acids Chains
Structures of the 20 common Amino Acids
Hydropathy
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Other Amino Acids and Amino Acid Derivatives
(GABA)
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Other Amino Acids and Amino Acid Derivatives
(21st amino acid)
(22nd amino acid)
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Ionization of Amino Acids
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Ionization of Amino Acids
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Ionization of Amino Acids
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Ionization of Amino Acids
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Ionization of Amino Acids
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Ionization of Amino Acids
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Peptide Bonds link Amino Acids in Proteins
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Peptide Bonds link Amino Acids in Proteins
Aspartame (aspartylphenylalanine methyl ester)
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Protein Purification Techniques
●Column chromatography /eluate/ ●HPLC
●Ion-exchange chromatography
●Gel-filtration chromatography
●Affinity chromatography
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Column chromatography /eluate/
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Analytical Techniques
●Polyacrylamide gel electrophoresis (PAGE) ●Mass spectrometry
●Electrospray mass spectrometry
●Matrix-assisted desorption ionization (MALDI)
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Polyacrylamide gel electrophoresis (PAGE)
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Matrix-assisted desorption ionization (MALDI)
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Matrix-assisted desorption ionization (MALDI)
MALDI-TOF
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Matrix-assisted desorption ionization (MALDI)
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Amino Acids Composition of Proteins
▪Peptide bonds of the protein are cleaved by acid hydrolysis using 6M HCl.
▪Method of amino acid analysis : treatment of the protein hydrolysate with PITC at pH 9.0 PTC-amino acid derivatives HPLC (column of fine silica beads+short hydrocarbon chains) Detection at 254 nm (peak absorbance of the PTC moiety)
▪1 picomole of a protein that contains about 200 residues
▪Glutamate + glutamine Glx or Z Aspartate + asparagine Asx or B
▪Side chain of Tryptophan is almost destroyed by acid hydrolysis
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Amino Acids Composition of Proteins
Acid hydrolysis
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Amino Acids Composition of Proteins(phenylisothiocyanate)
(phenylthiocarbomyl)
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Amino Acids Composition of Proteins
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Determing the Sequence of Amino Acid Residues
Edman degradation procedure
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Determing the Sequence of Amino Acid Residues
Entire procedure can be repeated serially using a sequenator
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Determing the Sequence of Amino Acid Residues
2-mercaptoethanol
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Protein Sequencing Strategies
▪Most proteins contain too many residues cleave peptide bonds proteases (trypsin, staphylococcus aureus V8 protease) or certain chemical reagents (BrCN)
▪BrCN reacts with Methionine C-terminal homoserine lactone Residues + new N-terminal residues
▪Trypsin carbonyl side of lysine and Arginine residues (positively charged side chains)
▪Staphylococcus aureus V8 protease carbonyl side of glutamate and Aspartate residues (negatively charged side chain)
▪Chymotrypsin carbonyl side of uncharged residues with aromatic or bulky hydrophobic side chains ( Phe, Tyr, Trp)
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Protein Sequencing Strategies
▪The amino acid sequence of a protein can be deduced from the sequence of nucleotides in the corresponding gene.
▪A sequence of three nucleotides specifies one amino acid.
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Comparisons of the Primary Structures of Proteins reveal Evolutionary Relationships
▪The protein cytochrome c (single polypeptide chain of about 104 residues) example of evolution at the molecular level
▪Protein sequences from distantly related species are similar enough proteins are homologous
▪Cytochrome c sequences of humans and chimpanzees are Identical.
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Comparisons of the Primary Structures of Proteins reveal Evolutionary Relationships