A heme tag for in vivo synthesis of artificial cytochromes.
A genetic approach is described here that enables the specific covalent attachment of heme via a short C-terminal peptide tag to an otherwise non-heme-binding protein. Covalent attachment of heme to the apo-protein is catalysed by the cytochrome c maturation system of Escherichia coli. While its original enzymatic activity is retained, the resulting heme-tagged protein is red, has peroxidase activity and is redox active. The presence or absence of a C-terminal histidine tag results in low-spin heme iron with six- or high-spin heme iron with five coordinate ligands, respectively. The heme tag can be used as a tool for the rational design of artificial c-type cytochromes and metalloenzymes, thereby overcoming previous limitations set by chemical approaches. Moreover, the tag allows direct visualisation of the red fusion protein during purification.
Braun
Martin
M
Institut für Mikrobiologie, ETH Hönggerberg, Wolfgang-Pauli-Str. 10, 8093, Zürich, Switzerland. mbraun@micro.biol.ethz.ch
Rubio
Inés García
IG
Thöny-Meyer
Linda
L
D000595:Amino Acid Sequence; D002352:Carrier Proteins / Q000378:metabolism; D002852:Chromatography, Ion Exchange; D045304:Cytochromes c / Q000096:biosynthesis*; D004926:Escherichia coli / Q000378:metabolism; D006418:Heme / Q000378:metabolism*; D058619:Maltose-Binding Proteins; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis*
D002352:Carrier Proteins; D058619:Maltose-Binding Proteins; D011993:Recombinant Fusion Proteins; D006418:Heme; D045304:Cytochromes c