The crystal structure of a heme oxygenase (HO) HugZ from Helicobacter pylori complexed with heme has been solved and refined at 1.8 Å resolution. HugZ is part of the iron acquisition mechanism of H. pylori, a major pathogen of human gastroenteric diseases. It is required for the adaptive colonization of H. pylori in hosts. Here, we report that HugZ is distinct from all other characterized HOs. It exists as a dimer in solution and in crystals, and the dimer adopts a split-barrel fold that is often found in FMN-binding proteins but has not been observed in hemoproteins. The heme is located at the intermonomer interface and is bound by both monomers. The heme iron is coordinated by the side chain of His 245 and an azide molecule when it is present in crystallization conditions. Experiments show that Arg 166 , which is involved in azide binding, is essential for HugZ enzymatic activity, whereas His 245 , surprisingly, is not, implying that HugZ has an enzymatic mechanism distinct from other HOs. The placement of the azide corroborates the observed ␥-meso specificity for the heme degradation reaction, in contrast to most known HOs that have ␣-meso specificity. We demonstrate through sequence and structural comparisons that HugZ belongs to a new heme-binding protein family with a split-barrel fold. Members of this family are widespread in pathogenic bacteria and may play important roles in the iron acquisition of these bacteria.One of the essential tasks for pathogenic bacteria to colonize mammalian hosts successfully is the acquisition of iron, an element required by all known life forms. Most bacteria require micromolar levels of iron for survival, but the element is usually not biologically available (1). The low concentration of free iron is caused by two factors: the low solubility (ϳ10 Ϫ18 M) of ferric iron at physiological pH, and the sequestration of ferrous iron into porphyrins and carrier molecules such as lactoferrin, transferrin, and ferritins. The latter is a mechanism employed by host cells to restrict microbe growth and to minimize the toxicity from superoxide anions and hydroxyl radicals generated by ferrous via Fenton reactions. To overcome iron shortages, bacteria have developed several mechanisms to import iron in different forms (1).Heme iron accounts for ϳ95% of iron in animal hosts (2), which suggests significant evolutionary pressure for pathogenic bacteria to prefer heme as an iron source, as has been demonstrated for Staphylococcus aureus (3). As a result, pathogenic bacteria have developed sophisticated systems to capture hemes, transfer them into bacterial cells, and degrade them to release free iron (4). Here, we report the 1.8 Å resolution crystal structure of HugZ (Hp0318), a heme oxygenase (HO) 4 essential for heme iron utilization by Helicobacter pylori. Wild-type H. pylori can use heme as a sole iron source (5), HugZ mutants, however, are unable to grow on heme iron (6).It has long been known that in eukaryotic cells, HOs, in conjunction with aerobic electron donors, carry out the degr...
