Cell modeling of heme deficiency using ferrochelatase mutations

摘要

Provided herein, is a means for development of ferrochelatase variants with improved tolerance towards N-methyl protoporphyrin. Also disclosed are cell assay systems utilizing the variants, as the variants would confer resistance to N-methyl protoporphyrin inhibition and thereby keep heme synthesis uninterrupted. The variants contain loop mutations that affect the NMPP-ferrochelatase interaction, and different degrees of NMPP tolerance are obtained with the introduction of loop mutations in wild-type ferrochelatase. Also disclosed is kinetic mechanism of inhibition of ferrochelatase by NMPP, using the disclosed variants whose mutations in the “porphyrin-interacting loop” motif weakened the potency of NMPP as an inhibitor.

主权项

1. A cellular model to determine physiological responses to heme deficiency comprising:
an isolated host cell expressing a ferrochelatase variant enzyme; wherein the ferrochelatase variant enzyme has a mutation in the ferrochelatase enzymatic active-site loop wherein there are no alterations outside of the ferrochelatase enzymatic active-site loop.