Expression and characterization of heme oxygenase, Hmx1, from Saccharomyces cerevisiae

Abstract

Heme oxygenase catalyzes the degradation of heme to biliverdin. The Saccharomyces cerevisiae heme oxygenase (Hmx1) gene minus the sequence coding for the last 23 amino acids that are the putative membrane-binding domain has been expressed in Escherichia coli. The truncated enzyme is obtained as a soluble and catalytically-active protein. The ferric form of the purified heme oxygenase has a Soret absorption maximum at 405 nm and its reduced ferrous-CO complex exhibits a typical Soret band at 420 nm and α/β bands at 568 and 532 nm. Titration with heme shows that Hmx1 is a heme binding protein, although heme is bound more slowly than it is to human heme oxygenase-1. Several reductants, including ascorbate, yeast cytochrome P450 reductase (CPR), human CPR, spinach ferredoxin/ferredoxin reductase, and putidaredoxin/putidaredoxin reductase, are able to provide electrons for biliverdin production by Hmx1. Of these, ascorbate appears to be the most effective reducing partner. Ascorbate-dependent heme oxidation by Hmx1 regiospecifically produces biliverdin IXα. Further characterization of the enzyme is in progress. Hmx1 is the first fungal heme oxygenase to be expressed to date, and its heme degradation activity suggests that Hmx1 plays a crucial role in iron and heme homeostasis in S. cerevisiae. (Supported by NIH R01 DK30297)