Effects of the size of magnetic particles of immobilized enzyme reactors on the digestion performance

Abstract

We applied immobilized enzyme reactors prepared with different sizes of magnetic particles into protein and proteome digestion. In addition, the influences of different sizes of the magnetic particles were studied on the reunion, enzyme efficiency and leakage sites. The experimental results showed that in comparison with the submicron magnetic particles, the amount of trypsin immobilized on the magnetic nanoparticles was 3. 5 times more than that of the submicron magnetic particles. However, the enzymatic efficiency was at the same level when the same amount of trypsin was used, and the reunion phenomenon was obviously improved when the size of the magnetic nanoparticles increased. Taking the immobilized enzyme reactor of 20 nm magnetic nanoparticles as an example, the digestion performance was further examined. The experimental results showed that rapid digestion could be achieved within 1 mm when the mass ratio of the trypsin and bovine serum albumin was 1:1. The peptide number of 0 missed cleavage site and the sequence coverage changed little after the protein was digested for 10 mm. It was concluded that the digestion efficiency of the immobilized enzyme reactor was much better than that of the in-solution digestion. When the immobilized enzyme reactors and the free trypsin were used for digestion, little differences of the leakage sites were found. Therefore, the immobilized enzyme reactors prepared with different sizes of magnetic particles can be applied in proteomic research for quick and efficient digestion.