Applicability of the alkylation chemistry for chemical C-terminal protein sequence analysis

Ne have evaluated the alkylation chemistry first described some years ago b y Boyd et al. which is now routinely applied in a commercial instrument. Ne have found that the low repetitive yields observed during these analyses a re due to the formation of a major side product when alkylating the C-termi nal thiohydantoin, This side product, resistant to the chemical cleavage me thods currently used, was characterized by NMR experiments in solution. We further demonstrate that chemical C-terminal sequence analysis of proteins using the alkylation chemistry is feasable with low picomole amounts of mat erial. High-sensitivity C-terminal sequencing allows a complementary approa ch by which a protein is first subjected to N-terminal Edman degradation fo llowed by C-terminal sequence analysis, limiting the amount of material nec essary for the characterization of the protein under study. This limited C- terminal sequence information is often sufficient to solve problems that ca nnot be solved by applying any other analytical method commonly used today.