In-gel protein N- and C-termini identification and its application for transgenic protein characterization

A new method for the determination of N‐ and C‐termini of a protein isolated in a polyacrylamide gel is introduced. In‐gel partial protein hydrolysis by hydrochloric acid is used to generate N‐ and C‐terminal peptides for identification. This new method is complementary to existing techniques. The application of the in‐gel protein termini identification technique to the characterization of the transgenic protein diacylglycerol acyltransferase (UrDGAT2A) purified from soybean seeds is also reported here. Both N‐ and C‐termini of UrDGAT2A were successfully identified and the N‐terminus was found to be blocked by acetylation. The analysis results of UrDGAT2A and two commercial proteins (bovine serum albumin (BSA) and alcohol dehydrogenase) are used to demonstrate the effectiveness of the method in identifying actual N‐ and C‐termini, terminal truncation and blocking. Copyright © 2010 John Wiley & Sons, Ltd.