Microwave assisted acid cleavage for denaturation and proteolysis of intact human adenovirus |
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Authors: | Fenselau Catherine Laine Olli Swatkoski Stephen |
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Institution: | Department of Chemistry & Biochemistry, University of Maryland, College Park MD 20742 USA. |
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Abstract: | Microwave assisted acid cleavage was applied directly to intact adenovirus type 5 to achieve denaturation and proteolysis in a single reaction. The speed of the digestion, coupled with the simplicity of MALDI analysis, allowed peptide products to be profiled in less than 5 min. Identification of peptides from a range of proteins by MALDI-TOF confirms that both denaturation and proteolysis were achieved using low concentrations of acetic acid (12.5%) and short incubations (1.5 to 2 min) at high temperatures (140° C). These conditions favor production of peptides that carry Asp on their C-termini. When this cleavage reaction is carried out in highly enriched H(2) (18)O, a single atom of (18)O is introduced site-specifically into the carboxyl terminal. The labeling reaction is applied to label reporter peptides from human adenovirus type 5 harvested from HeLa cells. Small peptide products of endogenous processing were also observed. |
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