BL/CH 401 Lecture #5

Protein Covalent Structure (Protein Primary Structure)

1. Peptide Bonds, Peptides and Proteins

2. Amino Acid Composition

3. Amino Acid Sequencing of Proteins and Peptides

General Approach to Determining the Amino Acid Sequence of a Protein:

1. Purify protein to homogeneity (See Lecture 6 & 7)

2. Determine AA composition (See #2 Above)

3. Reduce -S-S- bonds to -SH groups and block

4. Determine N- and C- termini

5. Cleave protein into peptides

6. Separate peptides

7. Determine amino acid sequence of the peptides

8. Use a second method to cleave the protein into peptides

9. Separate and sequence the second set of peptides

10. Deduce the entire amino acid sequence by identifying overlapping peptide sequences

After all peptides from one cleavage method have been sequenced, then the protein is cleaved by a second method and peptides separated and sequenced. So one sequences the protein twice. Overlapping sequences are identified so that the order of the peptides in the sequence can be deduced and consequently the entire amino acid sequence of the protein.

©Wilbur H. Campbell, 1995, 1996; wcampbel@mtu.edu

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