The HemQ coprohaem decarboxylase generates reactive oxygen species: implications for the evolution of classical haem biosynthesis

Abstract

Bacteria require a haem biosynthetic pathway for the assembly of a variety of protein complexes, including cytochromes, peroxidases, globins, and catalase. Haem is synthesised via a series of tetrapyrrole intermediates, including non-metallated porphyrins, such as protoporphyrin IX, which is well known to generate reactive oxygen species in the presence of light and oxygen. Staphylococcus aureus has an ancient haem biosynthetic pathway that proceeds via the formation of coproporphyrin III, a less reactive porphyrin. Here, we demonstrate, for the first time, that HemY of S. aureus is able to generate both protoporphyrin IX and coproporphyrin III, and that the terminal enzyme of this pathway, HemQ, can stimulate the generation of protoporphyrin IX (but not coproporphyrin III). Assays with hydrogen peroxide, horseradish peroxidase, superoxide dismutase, and catalase confirm that this stimulatory effect is mediated by superoxide. Structural modelling reveals that HemQ enzymes do not possess the structural attributes that are common to peroxidases that form compound I [FeIV==O]+, which taken together with the superoxide data leaves Fenton chemistry as a likely route for the superoxide-mediated stimulation of protoporphyrinogen IX oxidase activity of HemY. This generation of toxic free radicals could explain why HemQ enzymes have not been identified in organisms that synthesise haem via the classical protoporphyrin IX pathway. This work has implications for the divergent evolution of haem biosynthesis in ancestral microorganisms, and provides new structural and mechanistic insights into a recently discovered oxidative decarboxylase reaction.

Prokaryotic Heme Biosynthesis: Multiple Pathways to a Common Essential Product

“…Nevertheless, no Gram-positive organism that has the ability to synthesize protoporphyrin has currently been identified. The switch from the CPD pathway to the PPD pathway is an interesting one without explanation at present, but a recent report that ChdC stimulates CgoX to oxidize protoporphyrinogen to protoporphyrin may provide an interesting first clue (390).…”

Insights into the Active Site of Coproheme Decarboxylase from Listeria monocytogenes

“… 1 − 5 In fact, it catalyzes the decarboxylation of the two propionate groups at positions 2 and 4 of iron-coproporphyrin III (coproheme) to form heme b . Many recent publications have elucidated the physiological role of ChdC, 3 , 6 , 7 but its structure–function relationship is still not completely understood.…”

“…Coproheme decarboxylase (ChdC, formerly HemQ) is a key element in the coproporphyrin-dependent heme biosynthetic pathway of mainly monoderm, but also some diderm, archaea, and intermediate bacteria. − In fact, it catalyzes the decarboxylation of the two propionate groups at positions 2 and 4 of iron-coproporphyrin III (coproheme) to form heme b . Many recent publications have elucidated the physiological role of ChdC, ,, but its structure–function relationship is still not completely understood.…”

Epitope-specific immunity against Staphylococcus aureus coproporphyrinogen III oxidase

“…Indeed, identical aa sequences are deposited for S. aureus CgoX and protoporphyrinogen oxidase in public data banks like UniprotKB (Uniprot.org, compare Q2FXA5 and A0A0H3K8Y5). The CgoX-mediated generation of protoporphyrin IX, but not coproporphyrin III, is stimulated by heme-bound HemQ, which is mediated by superoxide 14 . TPI catalyses the reversible interconversion of the triose phosphate isomers dihydroxyacetone and D-glyceraldehyde 3-phosphate.…”

“…We here targeted two non-redundant S. aureus housekeeping proteins, coproporphyrinogen III oxidase (CgoX) and triose phosphate isomerase (TPI), which are essential for heme synthesis and glycolysis, respectively. Staphylococcal CgoX (EC: 1.3.3.15, also known as HemY) catalyses the oxidation of coproporphyrinogen III to coproporphyrin III 13 , 14 but can also oxidise protoporphyrinogen IX to protoporphyrin IX 15 . Indeed, identical aa sequences are deposited for S. aureus CgoX and protoporphyrinogen oxidase in public data banks like UniprotKB (Uniprot.org, compare Q2FXA5 and A0A0H3K8Y5).…”