Biomolecules Lecture_4_Proteins

Biomolecules Lecture_4_Proteins - Biomolecules and...

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Lecture 4 - Protein Sequencing, Evolution, and Purification Biomolecules and Metabolism
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Protein Structure 1. Primary 2. Secondary 3. Tertiary 4. Quartenary
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Amino Acid Sequence All amino sequences are read from the N terminal to C terminal end of the peptide bond Free carboxyl end (C-terminal) is written to the right Free amino end (N-terminal) is written to the left
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Amino Acid Code G - Glycine (Gly) P - Proline (Pro) A - Alanine (Ala) V - Valine (Val) L - Leucine (Leu I - Isoleucine (Ile) M - Methionine (Met) C - Cysteine (Cys) F - Phenylalanine (Phe) Y - Tyrosine (Tyr) W - Tryptophan (Trp) H - Histidine (His) K - Lysine (Lys) R - Arginine (Arg) Q - Glutamine (Gln) N - Asparagine (Asn) E - Glutamic Acid (Glu) D - Aspartic Acid (Asp) S - Serine (Ser) T - Threonine (Thr)
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Amino Acid Analysis
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Amino Acid Analysis
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Protein Sequencing: Why? 1. Allows comparison between normal and mutant proteins 2. Allows comparison between similar proteins in different species 3. Vital for prediction of protein 3D structures (an important aspect of rational drug design)
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Sequencing Steps 1. Purify the protein/peptide to homogeneity 2. Cleave all the disulphide bonds 3. Sequence: N-terminal sequencing C-terminal sequencing Edman sequencing
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N-Terminal Sequencing 1. FDNB label the N-terminal AA 2. Also known as ‘Sangers Reagent’ (2,4-dinitrofluorobenzene) 3. Hydrolyze polypeptide into its constituent AAs 4. N-terminal amino acid will be labeled with the DNB group (imparts a yellow colour) 5. Amino acids can be separated by electrophoresis or chromatography and labeled amino acid easily identified 6. Danysl chloride can also be used for this purpose (a fluorescent compound)
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N-Terminal Sequencing NH 2 - -COOH
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C-Terminal Sequencing 1. Hydrazine (H 2 N-NH
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This note was uploaded on 01/05/2010 for the course FSH BT taught by Professor Ianmarison during the Spring '09 term at Dublin City University.

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Biomolecules Lecture_4_Proteins - Biomolecules and...

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