This application note describes the N-terminal Sequence Analysis of peptide recovered from HPLC fractionated endoproteinase Lys-C digest using HP G1005A N-terminal Protein Sequencing System
This document details a procedure for analyzing peptide maps generated by HPLC fractionation of endoproteinase Lys-C digests. The specific example provided involves human apo-transferrin, demonstrating how peptides are sequenced through the C-terminal lysine residue using the HP G1005A N-terminal Protein Sequencing System.
The sample is dissolved in a buffer containing 8M urea, 100mM Tris.HCl at pH 9.0, and 10 mM DTT.
Endoproteinase Lys-C must be added to a ratio of 1:20 enzyme:substrate.
The separation is performed with a flowrate of 0.2 mL/min at a column temperature of 40°C.