Biochemical and Structural Characterization of Pseudomonas aeruginosa Bfd and FPR:  Ferredoxin NADP+ Reductase and Not Ferredoxin Is the Redox Partner of Heme Oxygenase under Iron-Starvation Conditions,

Wang, An; Zeng, Yuhong; Han, Huijong; Weeratunga, Saroja; Morgan, Bailey N.; Moënne‐Loccoz, Pierre; Schönbrunn, E.; Rivera, Mario

doi:10.1021/bi7013135

Cited by 40 publications

(84 citation statements)

References 68 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…4). Difficulties in releasing iron from the linear tetrapyrrole in vitro were reported for several HOs (Gisk et al 2012;Reniere et al 2010;Wang et al 2007;Wilks and Schmitt 1998). The efficiency of this final step in heme catalysis often appears to depend on the nature of the redox partner.…”

Section: Discussionmentioning

confidence: 99%

In vitro heme biotransformation by the HupZ enzyme from Group A streptococcus

et al. 2016

View full text Add to dashboard Cite

In Group A streptococcus (GAS), the metallorepressor MtsR regulates iron homeostasis. Here we describe a new MtsR-repressed gene, which we named hupZ (heme utilization protein). A recombinant HupZ protein was purified bound to heme from Escherichia coli grown in the presence of 5-aminolevulinic acid and iron. HupZ specifically binds heme with stoichiometry of 1:1. The addition of NADPH to heme-bound HupZ (in the presence of cytochrome P450 reductase, NADPH-regeneration system and catalase) triggered progressive decrease of the HupZ Soret band and the appearance of an absorption peak at 660 nm that was resistance to hydrolytic conditions. No spectral changes were observed when ferredoxin and ferredoxin reductase were used as redox partners. Differential spectroscopy with myoglobin or with the ferrous chelator, ferrozine, confirmed that carbon monoxide and free iron are produced during the reaction. ApoHupZ was crystallized as a homodimer with a split β-barrel conformation in each monomer comprising six β strands and three α helices. This structure resembles the split β-barrel domain shared by the members of a recently described family of heme degrading enzymes. However, HupZ is smaller and lacks key residues found in the proteins of the latter group. Phylogenetic analysis places HupZ on a clade separated from those for previously described heme oxygenases. In summary, we have identified a new GAS enzyme-containing split β-barrel and capable of heme biotransformation in vitro; to the best of our knowledge, this is the first enzyme among Streptococcus species with such activity.

show abstract

Section: Discussionmentioning

confidence: 99%

In vitro heme biotransformation by the HupZ enzyme from Group A streptococcus

et al. 2016

View full text Add to dashboard Cite

show abstract

“…For example, it also remains possible that FprA transfers electrons directly to CysI. It has been shown that P. aeruginosa FprA efficiently transfers electrons from NADPH to heme oxygenase (47). Annotated conserved domains within bacterial and chloroplast CysI proteins include "ferredoxin-like" sequences (48).…”

Section: Discussionmentioning

confidence: 99%

Role for Ferredoxin:NAD(P)H Oxidoreductase (FprA) in Sulfate Assimilation and Siderophore Biosynthesis in Pseudomonads

Lewis

Glassing

Harper

et al. 2013

Journal of Bacteriology

View full text Add to dashboard Cite

bPyridine-2,6-bis(thiocarboxylate) (PDTC), produced by certain pseudomonads, is a sulfur-containing siderophore that binds iron, as well as a wide range of transition metals, and it affects the net hydrolysis of the environmental contaminant carbon tetrachloride. The pathway of PDTC biosynthesis has not been defined. Here, we performed a transposon screen of Pseudomonas putida DSM 3601 to identify genes necessary for PDTC production (Pdt phenotype). Transposon insertions within genes for sulfate assimilation (cysD, cysNC, and cysG [cobA2]) dominated the collection of Pdt mutations. In addition, two insertions were within the gene for the LysR-type transcriptional activator FinR (PP1637). Phenotypic characterization indicated that finR mutants were cysteine bradytrophs. The Pdt phenotype of finR mutants could be complemented by the known target of FinR regulation, fprA (encoding ferredoxin:NADP ؉ oxidoreductase), or by Escherichia coli cysJI (encoding sulfite reductase). These data indicate that fprA is necessary for effective sulfate assimilation by P. putida and that the effect of finR mutation on PDTC production was due to deficient expression of fprA and sulfite reduction. fprA expression in both P. putida and P. aeruginosa was found to be regulated by FinR, but in a manner dependent upon reduced sulfur sources, implicating FinR in sulfur regulatory physiology. The genes and phenotypes identified in this study indicated a strong dependence upon intracellular reduced sulfur/ cysteine for PDTC biosynthesis and that pseudomonads utilize sulfite reduction enzymology distinct from that of E. coli and possibly similar to that of chloroplasts and other proteobacteria.

show abstract

“…Ferredoxin-NADP ? reductase from P. aeruginosa (pa-FPR) was expressed and purified as described before (Wang et al 2007). …”

Section: Production and Purification Of Recombinant Proteinsmentioning

confidence: 99%

“…The natural electron donor for PigA from P. aeruginosa PAO1 is pa-FPR (Wang et al 2007). Therefore, we performed additional experiments employing pa-FPR as electron donor instead of the artificial reductant system.…”

Section: Biochemical Characterization Of Pigasmentioning

confidence: 99%

Variable composition of heme oxygenases with different regiospecificities in Pseudomonas species

et al. 2012

View full text Add to dashboard Cite

Heme oxygenases (HO) degrade heme yielding iron, carbon monoxide and one of four possible biliverdin (BV) isomers. Pseudomonas aeruginosa PAO1 is thus far the only organism to contain two HOs with different regiospecificities: BphO and PigA. While BphO cleaves heme to exclusively yield BV IXα, PigA produces the BV isomers IXβ and IXδ. We bioinformatically identified putative HOs in diverse Pseudomonas strains, tested their enzymatic functionality and determined their regiospecificity. Surprisingly, even high amino acid sequence identities to the P. aeruginosa HOs were not sufficient to correctly predict the HO regiospecificity in all cases. Based on our results, Pseudomonas strains differ in their HO composition containing either BphO or PigA or both HO types. Concomitantly with the existence of bphO is the occurrence of at least one gene encoding a bacterial phytochrome implying that only BV IXα is the sufficient phytochrome chromophore. In contrast, pigA genes are organized in gene clusters associated with iron utilization implying a role of PigA in iron acquisition. However, at least in strains containing no PigA this function maybe fulfilled by BphO. Only a combination of homology searches and analyses of genetic environments is appropriate for a reliable prediction of the regiospecificity of Pseudomonas HOs.

show abstract

Biochemical and Structural Characterization of Pseudomonas aeruginosa Bfd and FPR: Ferredoxin NADP⁺ Reductase and Not Ferredoxin Is the Redox Partner of Heme Oxygenase under Iron-Starvation Conditions^,

Cited by 40 publications

References 68 publications

In vitro heme biotransformation by the HupZ enzyme from Group A streptococcus

In vitro heme biotransformation by the HupZ enzyme from Group A streptococcus

Role for Ferredoxin:NAD(P)H Oxidoreductase (FprA) in Sulfate Assimilation and Siderophore Biosynthesis in Pseudomonads

Variable composition of heme oxygenases with different regiospecificities in Pseudomonas species

Contact Info

Product

Resources

About