Tailoring recombinant lipases: keeping the His-tag favors esterification reactions, removing it favors hydrolysis reactions.

We determined the effect of the His-tag on the structure, activity, stability and immobilization of LipC12, a highly active lipase from a metagenomic library. We purified LipC12 with a N-terminal His-tag and then removed the tag using tobacco etch virus (TEV) protease. Circular dichroism analysis showed that the overall structure of LipC12 was largely unaffected by His-tag removal. The specific hydrolytic activities against natural and artificial substrates were significantly increased by the removal of the His-tag. On the other hand, His-tagged LipC12 was significantly more active and stable in the presence of polar organic solvents than untagged LipC12. The immobilization efficiency on Immobead 150 was 100% for both forms of LipC12 and protein desorption studies confirmed that the His-tag does not participate in the covalent binding of the enzyme. In the case of immobilized LipC12, the His-tag negatively influenced the hydrolytic activity, as it had for the free lipase, however, it positively influenced the esterification activity. These results raise the possibility of tailoring recombinant lipases for different applications, where the His-tag may be retained or removed, as appropriate for the desired activity.