Protein Sequencing Protocols

1. Front Matter

   Pages i-xiv

   PDF

2. Strategies for Handling Polypeptides on a Microscale

   • Bryan John Smith, Paul Tempst

   Pages 1-16

3. SDS Polyacrylamide Gel Electrophoresis for N-Terminal Protein Sequencing

   • Bryan John Smith

   Pages 17-24

4. Two-Dimensional Polyacrylamide Gel Electrophoresis for the Separation of Proteins for Chemical Characterization

   • Michael J. Dunn

   Pages 25-36

5. Electroblotting of Proteins from Polyacrylamide Gels for Chemical Characterization

   • Michael J. Dunn

   Pages 37-42

6. Enzymatic Cleavage of Polypeptides

   • Bryan John Smith, Colin Wheeler

   Pages 43-55

7. Chemical Cleavage of Polypeptides

   • Bryan John Smith

   Pages 57-72

8. Preparation of Glycopeptides

   • Chris W. Sutton, Jacqui A. O’Neill

   Pages 73-79

9. Selective Isolation of the Carboxy Terminal Peptide from a Protein

   • Tomoko Hayashi Akiyama, Tatsuru Sasagawa

   Pages 81-83

10. Selective Isolation of the Amino-Terminal Peptide from α-Amino Blocked Protein

    • Tomoko Hayashi Akiyama, Tatsuru Sasagawa

    Pages 85-89

11. Analytical and Micropreparative Capillary Electrophoresis of Peptides

    • Alan J. Smith

    Pages 91-99

12. Reverse-Phase HPLC Purification of Peptides from Natural Sources for Structural Analysis

    • Chris Shaw

    Pages 101-107

13. High-Performance Liquid Chromatography On-Line Derivative Spectroscopy for the Characterization of Peptides with Aromatic Amino Acid Residues

    • Christoph W. Turck

    Pages 109-117

14. Hydrolysis of Samples for Amino Acid Analysis

    • Ian Davidson

    Pages 119-129

15. Amino Acid Analysis

    • G. Brent Irvine

    Pages 131-138

16. Postcolumn Amino Acid Analysis

    • Alan J. Smith

    Pages 139-146

17. Fast-Atom Bombardment and Electrospray Mass Spectrometry of Peptides, Proteins, and Glycoproteins

    • Fiona M. Greer, Howard R. Morris

    Pages 147-163

18. Peptide Mass Fingerprinting Using MALDI-TOF Mass Spectrometry

    • Darryl J. C. Pappin

    Pages 165-173

19. N-Terminal Protein Ladder Sequencing

    • Rong Wang, Brian T. Chait

    Pages 175-182

20. The Dansyl-Edman Method for Manual Peptide Sequencing

    • John M. Walker

    Pages 183-187

If the development of techniques for the labeling of a polypeptide- terminus and for repetitive N-terminal sequencing mark the beginning of the science of protein structure determination, then the field has just about reached its half century. In more recent times, recombinant DNA techniques have pro­ vided powerful means by which to obtain long protein sequences (by theoreti­ cal translation from nucleic acid sequences), but rather than replacing the direct, chemical, protein sequencing approach, they have instead added further impe­ tus to the drive towards a better understanding of posttranslational processing and modification events, as well as identification of novel proteins. Recent years have also seen the advent of "biopharmaceuticals" (i. e. , pharmaceutical products that are proteins, such as monoclonal antibodies), and this has meant that protein sequencing has found an important new application as a quality control tool. Over the decades of protein sequencing many new techniques have been introduced, with the basic aim of generating more information from less material. Some techniques have come and gone, but others have been with us for many years. Edman chemistry is perhaps the best example of the latter class, with its basic principles still being applied today in the newest protein sequencer design. Methods for cleaving and modifying peptides are also long­ standing, having been adapted over the years to suit progressively smaller amounts of sample.

"This methodological guide can be recommended to all who are concerned with protein characterization or peptide mapping."-Reviewed by R. Moravec, Biologia Plantarum

• Celltech Therapeutics, Slough, UK

  Bryan John Smith

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