A heme tag for in vivo synthesis of artificial cytochromes |
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Authors: | Martin Braun Inés García Rubio Linda Thöny-Meyer |
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Institution: | (1) Institut für Mikrobiologie, ETH Hönggerberg, Wolfgang-Pauli-Str. 10, 8093 Zürich, Switzerland;(2) Laboratorium für Physikalische Chemie, Eidgenössische Technische Hochschule, Wolfgang-Pauli-Str. 10, 8093 Zürich, Switzerland |
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Abstract: | 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. |
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