The genetic strategy used to insert the His9 tag into MBH.
A, the tag is inserted at the C terminus of mbhN (yielding P. furiosus strain MW0403) where PF1437 is the gene immediately downstream of the MBH operon. B, the tag is inserted at the N terminus of mbhJ (yielding P. furiosus strain MW0414). UFR, upstream flanking region (1 kb) of the MBH operon; DFR, downstream flanking region (1 kb) of the MBH operon; pyrF, selectable marker; Pgdh and Pslp, promoters for the gene encoding glutamate dehydrogenase and the S-layer protein of P. furiosus, respectively. The affinity tag was a His9 tag.