Ni-modified magnetic nanoparticles for affinity purification of His-tagged proteins from the complex matrix of the silkworm fat body.

Purification of recombinant proteins is often a challenging matter because high purity and high recovery are desired. If the expressed recombinant protein is also in a complex matrix, such as from the silkworm expression system, purification becomes more challenging. Even if purification from the silkworm expression system is troublesome, it benefits from a high capacity for the production of recombinant proteins. In this study, magnetic nanoparticles (MNPs) were investigated as a suitable tool for the purification of proteins from the complex matrix of the silkworm fat body. The MNPs were modified with nickel so that they have an affinity for His-tagged proteins, as the MNP purification protocol itself does not need special equipment except for a magnet. Among the three different kinds of investigated MNPs, MNPs with sizes of 100 nm to 200 nm and approximately 20 nm-thick nickel shells were the most suitable for our purpose. With them, the total protein amount was reduced by up to at least approximately 77.7%, with a protein recovery of around 50.8% from the silkworm fat body. The minimum binding capacity was estimated to be 83.3 µg protein/mg MNP. Therefore, these MNPs are a promising tool as a purification pretreatment of complex sample matrices.