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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
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)
General Structure of Amino Acids
General Structure of Amino Acids
Zwitterions
Chiral OrAsymmetric
General Structure of Amino Acids
Ball-and-stick model ofserine
General Structure of Amino AcidsStereoisomers (enantiomers)
General Structure of Amino Acids
The RS system
General Structure of Amino Acids
The RS system
Clockwise R configuration
Counterclockwise S configuration
Structures of the 20 common Amino Acids
A. Aliphatic R Groups
A. Aliphatic R Groups
Structures of the 20 common Amino Acids
A. Aliphatic R Groups
Structures of the 20 common Amino Acids
Pyrrolidine ring
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
B. Aromatic R Groups
Structures of the 20 common Amino Acids
(Benzyl) (Phenol) (Indol)(260 nm) (280 nm) (280 nm)
C. Sulfur-Containing R Groups
Structures of the 20 common Amino Acids
Disulfide bond
C. Sulfur-Containing R Groups
Structures of the 20 common Amino Acids
Nonpolar Methyl thioether
Sulfhydryl
D. Side Chains with Alcohol Groups
Structures of the 20 common Amino Acids
α- carbon
β- carbon
Stereoisomers
E. Basic R Groups
Structures of the 20 common Amino Acids
Imidazole ring
Guanidinium ion
F. Acidic R Groups and Their Amide Derivatives
Structures of the 20 common Amino Acids
Amide
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.
G. The Hydrophobicity of Amino Acids Chains
Structures of the 20 common Amino Acids
Hydropathy
Other Amino Acids and Amino Acid Derivatives
(GABA)
Other Amino Acids and Amino Acid Derivatives
(21st amino acid)
(22nd amino acid)
Ionization of Amino Acids
Ionization of Amino Acids
Ionization of Amino Acids
Ionization of Amino Acids
Ionization of Amino Acids
Ionization of Amino Acids
Peptide Bonds link Amino Acids in Proteins
Peptide Bonds link Amino Acids in Proteins
Aspartame (aspartylphenylalanine methyl ester)
Protein Purification Techniques
●Column chromatography /eluate/ ●HPLC
●Ion-exchange chromatography
●Gel-filtration chromatography
●Affinity chromatography
Column chromatography /eluate/
Analytical Techniques
●Polyacrylamide gel electrophoresis (PAGE) ●Mass spectrometry
●Electrospray mass spectrometry
●Matrix-assisted desorption ionization (MALDI)
Polyacrylamide gel electrophoresis (PAGE)
Matrix-assisted desorption ionization (MALDI)
Matrix-assisted desorption ionization (MALDI)
MALDI-TOF
Matrix-assisted desorption ionization (MALDI)
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
Amino Acids Composition of Proteins
Acid hydrolysis
Amino Acids Composition of Proteins(phenylisothiocyanate)
(phenylthiocarbomyl)
Amino Acids Composition of Proteins
Determing the Sequence of Amino Acid Residues
Edman degradation procedure
Determing the Sequence of Amino Acid Residues
Entire procedure can be repeated serially using a sequenator
Determing the Sequence of Amino Acid Residues
2-mercaptoethanol
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)
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.
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.
Comparisons of the Primary Structures of Proteins reveal Evolutionary Relationships
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