N-Terminal Sequencing (Amino Terminal Sequencing / Edman Sequencing)
N-Terminal (amino-terminal) sequencing by automated Edman chemistry, also known as automated Gas Phase Sequencing (GPS), is used in protein characterisation to determine the amino-terminus of the peptide or protein. It is used throughout all stages of drug discovery, or to demonstrate comparability and consistency between batches for release during manufacturing. It is also used during discovery for de novo sequencing of new novel proteins and is available as part of SGS M-Scan’s full protein characterization package and for antibody analysis.
N-Terminal sequencing can be undertaken on samples in solution or from SDS-PAGE separated proteins blotted on to PVDF membranes. It can be used for the analysis of possible truncations of the amino-terminus, as well as to assess the level of chemically blocked termini which generally prevents full sequencing. It may be possible to remove the blocking group prior to sequencing.
Automated Edman chemistry sequentially breaks down a peptide/protein into its constituent amino acids from the N-terminus of the sample. The produced amino acid is derivatised and separated by RP-HPLC and visualised by UV detection for each cycle of Edman chemistry. The amino acids are quantified by comparison to a standard mixture.
N-Terminal sequencing is a key part of the ICH Q6B guidelines for characterization and confirmation of biopharmaceuticals in support of new marketing applications.