Findings

Crystal Formation

After performing crystal trials, the His-Tag VP8 protein was successfully crystallized. The previously identified substrate, LNDF1 (C32H55N1O25), was used to soak the crystal for binding analysis. However, when the bat rotavirus His-Tag VP8 protein was introduced to the crystals, no observable interaction or binding occurred. This suggested that the spike protein VP8 failed to interact with the substrate in its native form.

Modifying the Protein: GST Tag Addition and Cleavage

Given the lack of interaction between the protein VP8 and the LNDF1 crystals, the next step involved modifying the VP8 protein by adding a GST tag. This modification was aimed at enhancing the protein's binding affinity to the substrate. After the GST tag was introduced, it was subsequently cleaved off to restore the protein's original structure and functionality, potentially facilitating more effective substrate interaction.