Trypsin immobilization on silica beads modified by squamous polymer for ultra fast and highly efficient proteome digestion

Abstract

Currently, the shotgun based strategy has been widely applied in proteomic research. In this strategy, protein identification relied on the identification of the corresponding proteolytic peptides. Therefore, rapid and efficient protein digestion is crucial for accurate protein identification and characterization. Even though traditional free protein digestion in solution has been widely adopted, it had a few inherent disadvantages including long incubation time, incomplete digestion and non-reusability of the protease. In this work, we developed a new type of trypsin immobilized on squamous polymer modified silica bead (SPMSB) for ultra fast and highly efficient protein digestion. The squamous polymer coated silica beads were prepared by surface initiated atom transfer radical polymerization (SI-ATRP), which leaded to surface confined growth of non-crosslinked polymer chains on the sufface of the silica beads for trypsin immobilization. The digestion efficiency of the obtained SPMSB-trypsin was evaluated using both standard proteins and complex protein extracts obtained from E. coil. Highly efficient digestion was achieved in only 1 - 2 mm digestion. Furthermore, the SPMSB-Trypsin exhibited both good stability and excellent recovery, therefore can be applied in proteomic research in the future.