Oxidation of Heme to β- and δ-Biliverdin by<i>Pseudomonas a</i><i>eruginosa</i>Heme Oxygenase as a Consequence of an Unusual Seating of the Heme

Caignan, Gregori A.; Deshmukh, Rahul; Wilks, Angela; Zeng, Yuhong; Huang, Huaxiong; Moënne‐Loccoz, Pierre; Bunce, Richard A.; Eastman, Margaret A.; Rivera, Mario

doi:10.1021/ja0274960

Cited by 96 publications

(214 citation statements)

References 59 publications

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“…The low activation of entropy of the wild-type pa-HO suggests that the transition state is highly ordered, and this stabilization could be provided in the form of a protein-protein interaction, which is consistent with the gain in entropy observed for the pa-HO-mutants and BphO. However, an exact interpretation of these effects will require a more extensive study with a larger set of mutants at these and other positions in and around the heme pocket of both PhuS and pa-HO.Previous observations suggested that at neutral pH the heme-PhuS and heme-pa-HO complexes are respectively (15,19). Therefore, changes in both the spin-state and axial heme coordination must occur during the transfer reaction.…”

supporting

confidence: 77%

“…The increase in the activation energy for heme transfer from holo-PhuS to the pa-HO mutants relative to the native protein is most likely due to the loss of favorable binding or interaction between PhuS and pa-HO. The pa-HO mutants introduced heme propionate interactions required for α-regioselective specificity (pa-HO-N19K/F117Y) as in other characterized HO enzymes and additionally removed residues that stabilize the δ-regioselective heme seating (pa-HO-N19K/ F117Y/K132A and pa-HO-N19K/F117Y/K34A) (19). In addition, these amino acid replacements are located on the heme binding face of pa-HO and, as well as stabilizing the bound heme, are located such that in the apo-pa-HO they may provide additional electrostatic or hydrogen-bonding interactions with the holo-PhuS (22).…”

Section: Discussionmentioning

confidence: 99%

“…These mutations have previously been shown to destabilize the heme within the protein such that it is in dynamic in-plane rotation between two seatings that yield an altered isomer pattern than that of the wild-type protein (19,22). Therefore we would predict that such a destabilization would have an effect on the rate of heme transfer.…”

Section: Heme Transfer Experimentsmentioning

confidence: 98%

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The Mechanism of Heme Transfer from the Cytoplasmic Heme Binding Protein PhuS to the δ-Regioselective Heme Oxygenase of Pseudomonas aeruginosa

Bhakta

Wilks

2006

Biochemistry

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The opportunistic pathogen Pseudomonas aeruginosa has evolved two outer membrane receptor mediated uptake systems (encoded by the phu and has operons) by which it can utilize the hosts heme and hemeproteins as a source of iron. PhuS is a cytoplasmic heme binding protein encoded within the phu operon, and has previously been shown to function in the trafficking of heme to the ironregulated heme oxygenase (pa-HO). While the heme association rate for PhuS was similar to that of myoglobin, a markedly higher rate of heme dissociation (∼10 5 s −1 ) was observed, in keeping with a function in heme-trafficking. Additionally, the transfer of heme from PhuS to pa-HO was shown to be specific and unidirectional when compared to transfer to the non-iron regulated heme oxygenase (BphO), in which heme distribution between the two proteins merely reflects their relative intrinsic affinities for heme. Furthermore, the rate of transfer of heme from holo-PhuS to pa-HO of 0.11 ± 0.01 s −1 is 30-fold faster than that to apo-myoglobin, despite the significant higher binding affinity of apo-myoglobin for heme (k H =1.3 × 10 −8 μM) than that of PhuS (0.2 μM). This data suggests that heme transfer to pa-HO is independent of heme affinity and is consistent with temperature dependence studies which indicate the reaction is driven by a negative entropic contribution, typical of an ordered transition state, and supports the notion that heme transfer from PhuS to pa-HO is mediated via a specific protein-protein interaction. In addition, pH studies, and reactions conducted in the presence of cyanide, suggest the involvement of spin transition during the heme transfer process, whereby the heme undergoes spin change from 6-c LS to 6-c HS either in PhuS or pa-HO. Based on the magnitudes of the activation parameters obtained in the presence of cyanide, whereby both complexes are maintained in a 6-c LS state, and the biphasic kinetics of heme transfer from holo-PhuS to pa-HO-wt, supports the notion that the spin-state crossover occur within holo-PhuS prior to heme transfer step. Alternatively, the lack of the biphasic kinetic with pa-HO-G125V, 6-c LS, and with comparable rate of heme transfer as pa-HO-is supportive of mechanism in which the spin-change could occur within pa-HO. The present data suggests either or both of the two pathways proposed for heme transfer may occur under the present experimental conditions. The dissection of which pathway is physiologically relevant is the focus of ongoing studies.Heme, a cofactor of proteins involved in a variety of biological processes such as oxygen transport and storage, oxygenation reactions, electron transfer and transcriptional regulation is also a redox-reactive, hydrophobic iron chelate that readily associates with membranes, and is toxic to cells due to its ability to generate reactive oxygen species. Therefore, aerobic organisms have developed strategies to protect themselves from the harmful effects of "free" heme by sequestering it within specific proteins (1,2). While hemeproteins se...

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supporting

confidence: 77%

Section: Discussionmentioning

confidence: 99%

Section: Heme Transfer Experimentsmentioning

confidence: 98%

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The Mechanism of Heme Transfer from the Cytoplasmic Heme Binding Protein PhuS to the δ-Regioselective Heme Oxygenase of Pseudomonas aeruginosa

Bhakta

Wilks

2006

Biochemistry

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“…10 The HOs identified for a number of pathogeneic bacteria 11-13 appear to have as their major role the "mining" of iron to infect a host. 3, 5 Two such HOs of interest here are Neisseria meningitidis, NmHO (also called HemO 12,14,15 ) and Pseudomonas aeruginosa, PaHO 13,16,17 . A common mechanism, worked out on the mammalian HOs, appears operative in the various HOs.…”

Section: Introductionmentioning

confidence: 99%

¹H NMR Study of the Magnetic Properties and Electronic Structure of the Hydroxide Complex of Substrate-bound Heme Oxygenase from Neisseria Meningitidis: Influence of the Axial Water Deprotonation on the Distal H-bond Network

Liu

Zhang

et al. 2006

J. Am. Chem. Soc.

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The substrate and active site residues of the low-spin hydroxide complex of the protohemin complex Neisseria meningitidis heme oxygenase, HO, NmHO, have been assigned by saturation transfer between the hydroxide and previously characterized aquo complex. The available dipolar shifts allowed the quantitation of both the orientation and anisotropy of the paramagnetic susceptibility tensor. The resulting positive sign, and reduced magnitude of the axial anisotropy relative to the cyanide complex, dictate that the orbital ground state is the conventional '

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“…1). The unusual production of ␤-and ␦-BV was shown to be due to an unusual seating of the substrate heme in the active site pocket of PigA (9). The purpose for the production of these BV isomers and their fate in P. aeruginosa, however, is currently unknown.…”

mentioning

confidence: 99%

The Heme Oxygenase(s)-Phytochrome System of Pseudomonas aeruginosa

Wegele

Tasler

Zeng

et al. 2004

Journal of Biological Chemistry

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136

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For many pathogenic bacteria like Pseudomonas aeruginosa heme is an essential source of iron. After uptake, the heme molecule is degraded by heme oxygenases to yield iron, carbon monoxide, and biliverdin. The heme oxygenase PigA is only induced under ironlimiting conditions and produces the unusual biliverdin isomers IX␤ and IX␦. The gene for a second putative heme oxygenase in P. aeruginosa, bphO, occurs in an operon with the gene bphP encoding a bacterial phytochrome. Here we provide biochemical evidence that bphO encodes for a second heme oxygenase in P. aeruginosa. HPLC, 1 H, and 13 C NMR studies indicate that BphO is a "classic" heme oxygenase in that it produces biliverdin IX␣. The data also suggest that the overall fold of BphO is likely to be the same as that reported for other ␣-hydroxylating heme oxygenases. Recombinant BphO was shown to prefer ferredoxins or ascorbate as a source of reducing equivalents in vitro and the ratelimiting step for the oxidation of heme to biliverdin is the release of product. In eukaryotes, the release of biliverdin is driven by biliverdin reductase, the subsequent enzyme in heme catabolism. Because P. aeruginosa lacks a biliverdin reductase homologue, data are presented indicating an involvement of the bacterial phytochrome BphP in biliverdin release from BphO and possibly from PigA.

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Oxidation of Heme to β- and δ-Biliverdin byPseudomonas aeruginosaHeme Oxygenase as a Consequence of an Unusual Seating of the Heme

Cited by 96 publications

References 59 publications

The Mechanism of Heme Transfer from the Cytoplasmic Heme Binding Protein PhuS to the δ-Regioselective Heme Oxygenase of Pseudomonas aeruginosa

The Mechanism of Heme Transfer from the Cytoplasmic Heme Binding Protein PhuS to the δ-Regioselective Heme Oxygenase of Pseudomonas aeruginosa

¹H NMR Study of the Magnetic Properties and Electronic Structure of the Hydroxide Complex of Substrate-bound Heme Oxygenase from Neisseria Meningitidis: Influence of the Axial Water Deprotonation on the Distal H-bond Network

The Heme Oxygenase(s)-Phytochrome System of Pseudomonas aeruginosa

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Oxidation of Heme to β- and δ-Biliverdin byPseudomonas aeruginosaHeme Oxygenase as a Consequence of an Unusual Seating of the Heme

Cited by 96 publications

References 59 publications

The Mechanism of Heme Transfer from the Cytoplasmic Heme Binding Protein PhuS to the δ-Regioselective Heme Oxygenase of Pseudomonas aeruginosa

The Mechanism of Heme Transfer from the Cytoplasmic Heme Binding Protein PhuS to the δ-Regioselective Heme Oxygenase of Pseudomonas aeruginosa

1H NMR Study of the Magnetic Properties and Electronic Structure of the Hydroxide Complex of Substrate-bound Heme Oxygenase from Neisseria Meningitidis: Influence of the Axial Water Deprotonation on the Distal H-bond Network

The Heme Oxygenase(s)-Phytochrome System of Pseudomonas aeruginosa

Contact Info

Product

Resources

About

¹H NMR Study of the Magnetic Properties and Electronic Structure of the Hydroxide Complex of Substrate-bound Heme Oxygenase from Neisseria Meningitidis: Influence of the Axial Water Deprotonation on the Distal H-bond Network