Ch. 5 - Protein TechniquesWorksheetSee all chapters
All Chapters
Ch. 1 - Introduction to Biochemistry
Ch. 2 - Water
Ch. 3 - Amino Acids
Ch. 4 - Protein Structure
Ch. 5 - Protein Techniques
Ch. 6 - Enzymes and Enzyme Kinetics
Ch. 7 - Enzyme Inhibition and Regulation
Ch. 8 - Protein Function
Ch. 9 - Carbohydrates
Ch. 10 - Lipids
Ch. 11 - Biological Membranes and Transport
Ch. 12 - Biosignaling
Clutch Review 1: Nucleic Acids, Lipids, & Membranes
Clutch Review 2: Biosignaling, Glycolysis, Gluconeogenesis, & PP-Pathway
Clutch Review 3: Pyruvate & Fatty Acid Oxidation, Citric Acid Cycle, & Glycogen Metabolism
Clutch Review 4: Amino Acid Oxidation, Oxidative Phosphorylation, & Photophosphorylation
Protein Purification
Protein Extraction
Differential Centrifugation
Salting Out
Column Chromatography
Ion-Exchange Chromatography
Anion-Exchange Chromatography
Size Exclusion Chromatography
Affinity Chromatography
Specific Activity
Native Gel Electrophoresis
SDS-PAGE Strategies
Isoelectric Focusing
Diagonal Electrophoresis
Mass Spectrometry
Mass Spectrum
Tandem Mass Spectrometry
Peptide Mass Fingerprinting
Overview of Direct Protein Sequencing
Amino Acid Hydrolysis
Chemical Cleavage of Bonds
Edman Degradation
Edman Degradation Sequenator and Sequencing Data Analysis
Edman Degradation Reaction Efficiency
Ordering Cleaved Fragments
Strategy for Ordering Cleaved Fragments
Indirect Protein Sequencing Via Geneomic Analyses

Concept #1: Mass Spectra Can Reveal Primary Protein Structure

Practice: Use the mass spectrum below to determine the sequence of the peptide.

Concept #2: Y Ions and B Ions

Practice: Upon fragmentation of a peptide bond during mass spectrometry, what ions can be detected on the spectrum?

Practice: Use the mass spectrum below & the indicated y-ion peaks (red) to reveal the sequence of the peptide.

Practice: In your mass-spectrometry of a pure protein with an m/z of 1,582, you found peaks of y ions with the following m/z ratios of 1,582, 1396 and 1283. The mass in Daltons for the possible relevant amino acids are provided: Y (163), N (114), W (186), D (115), G (57), L (113) and M (131). From this data, it is obvious that the C-terminal amino acid residue of the 1,582 fragment is:

Practice: Use the mass spectrum below & the provided chart with amino acid masses to determine the sequence of a hexapeptide (6 amino acid residues). In the mass spectrum, y ion peaks are indicated with “y” while b ion peaks are indicated with “b.” The N-terminal residue is given as Leu and the C-terminal residue is given as Lys. Determine the remaining amino acid sequence using either the y ions or the b ions.