Enhanced responses in matrix-assisted laser desorption/ionization mass spectrometry of peptides derivatized with arginine via a C-terminal oxazolone

We have developed a novel method for enhancing the response of a peptide in matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) by activating the C-terminal carboxyl group through an oxazolone with which is coupled an amine containing a functional group to help ionize the peptide. The reactions consist of dehydration with acetic anhydride to give an oxazolone, followed by aminolysis with an appropriate amino acid derivative such as arginine methyl ester. The MALDI signal of Ac-Tyr-Gly-Gly-Phe-Leu-Arg-OMe, thus converted from leucine-enkephalin, was detected while completely excluding the responses of arginine-deficient peptides coexisting in the reaction mixture. Some less intense peaks corresponding to a few sequential degradation products, also terminated with the arginine derivative, were also observed. The side-chain groups potentially that are reactive were conveniently protected by acetylation simultaneous with the C-terminal activation, and those that remained unprotected were reduced to virtually negligible proportions when the reaction was conducted in a peptide solution of concentration less than 1 mM. The greatly increased responses of such arginine-terminated peptides could possibly be exploited to discern the C-terminal tryptic peptide of a protein that is otherwise almost insensitive to MALDI-MS in general. The simplicity of the post-source decay spectrum of enkephalin derivatized by arginine methyl ester characteristically accentuated z- and b-type ions, and this should facilitate sequencing of such derivatized peptides. Remaining problems with practical applications of this approach are discussed. Copyright (C) 2004 John Wiley Sons, Ltd.