Accumulation and drainage of hemin in the red cell membrane

Shaklai, Nurith; Shviro, Yocheved; Rabizadeh, Esther; Kirschner-Zilber, I.

doi:10.1016/0005-2736(85)90106-3

Cited by 93 publications

(43 citation statements)

References 27 publications

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“…In this regulatory pathway, the direct consequence of decreased oxygen levels is the depletion of heme and sterols, which causes an increase in membrane fluidity. This step is supported by our result that chemicals that decrease membrane fluidity (DMSO and glycerol) block the hypoxic induction of PAU transcription and that the levels of both ergosterol (Lees et al 1979;Berg et al 2002;Abe et al 2009) and heme (Kirschner-Zilber et al 1982;Shviro et al 1982;Ginsburg and Demel 1984;Shaklai et al 1985;Light and Olson 1990;Balla et al 1991;Schmitt et al 1993) affect membrane fluidity.…”

Section: Discussionmentioning

confidence: 53%

The Hog1 Mitogen-Activated Protein Kinase Mediates a Hypoxic Response inSaccharomyces cerevisiae

2011

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We have studied hypoxic induction of transcription by studying the seripauperin (PAU ) genes of Saccharomyces cerevisiae. Previous studies showed that PAU induction requires the depletion of heme and is dependent upon the transcription factor Upc2. We have now identified additional factors required for PAU induction during hypoxia, including Hog1, a mitogen-activated protein kinase (MAPK) whose signaling pathway originates at the membrane. Our results have led to a model in which heme and ergosterol depletion alters membrane fluidity, thereby activating Hog1 for hypoxic induction. Hypoxic activation of Hog1 is distinct from its previously characterized response to osmotic stress, as the two conditions cause different transcriptional consequences. Furthermore, Hog1-dependent hypoxic activation is independent of the S. cerevisiae general stress response. In addition to Hog1, specific components of the SAGA coactivator complex, including Spt20 and Sgf73, are also required for PAU induction. Interestingly, the mammalian ortholog of Spt20, p38IP, has been previously shown to interact with the mammalian ortholog of Hog1, p38. Taken together, our results have uncovered a previously unknown hypoxic-response pathway that may be conserved throughout eukaryotes.

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Section: Discussionmentioning

confidence: 53%

The Hog1 Mitogen-Activated Protein Kinase Mediates a Hypoxic Response inSaccharomyces cerevisiae

2011

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“…The heme concentration was estimated from absorbance at 385 nm (⑀ m M ϭ 58,400) in 100 mM NaOH (16) and adjusted to 1.0 mM. This stock reagent was stored in the dark on ice and used within 24 h.…”

Section: Methodsmentioning

confidence: 99%

Clotrimazole Binds to Heme and Enhances Heme-dependent Hemolysis

Huy

Kamei

Yamamoto

et al. 2002

Journal of Biological Chemistry

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Two recent studies have demonstrated that clotrimazole, a potent antifungal agent, inhibits the growth of chloroquine-resistant strains of the malaria parasite, Plasmodium falciparum, in vitro. We explored the mechanism of antimalarial activity of clotrimazole in relation to hemoglobin catabolism in the malaria parasite. Because free heme produced from hemoglobin catabolism is highly toxic to the malaria parasite, the parasite protects itself by polymerizing heme into insoluble nontoxic hemozoin or by decomposing heme coupled to reduced glutathione. We have shown that clotrimazole has a high binding affinity for heme in aqueous 40% dimethyl sulfoxide solution (association equilibrium constant: K a ‫؍‬ 6.54 ؋ 10 8 M ؊2 ). Even in water, clotrimazole formed a stable and soluble complex with heme and suppressed its aggregation. The results of optical absorption spectroscopy and electron spin resonance spectroscopy revealed that the heme-clotrimazole complex assumes a ferric low spin state (S ‫؍‬ 1 ⁄2), having two nitrogenous ligands derived from the imidazole moieties of two clotrimazole molecules. Furthermore, we found that the formation of heme-clotrimazole complexes protects heme from degradation by reduced glutathione, and the complex damages the cell membrane more than free heme. The results described herein indicate that the antimalarial activity of clotrimazole might be due to a disturbance of hemoglobin catabolism in the malaria parasite.

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“…After cooling on ice for 5 min, the solution was centrifuged to remove insoluble particulates and the supernatant transferred to a fresh tube. The concentration of the heme solution was determined spectrophotometrically in 0.1 N NaOH using the extinction coefficient 58.4 mM −1 cm −1 at 385 nm (29). Heme standards prepared in 100 mM potassium phosphate pH 7.4 and labeled samples were boiled 3 min, placed on ice 5 min to cool, and then centrifuged 10 min at 15,000 rpm to pellet denatured protein.…”

Section: Cyp2e1 Integritymentioning

confidence: 99%

CYP2E1 active site residues in substrate recognition sequence 5 identified by photoaffinity labeling and homology modeling

Collom¹,

Jamakhandi²,

Tackett³

et al. 2007

Archives of Biochemistry and Biophysics

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Despite its biological importance, our knowledge of active site structure and relevance of critical amino acids in CYP2E1 catalytic processes remain limited. In this study, we identified CYP2E1 active site residues using photoaffinity labeling with 7-azido-4-methylcoumarin (AzMC) coupled with a CYP2E1 homology model. In the absence of light, AzMC was an effective competitor against substrate p-nitrophenol oxidation by CYP2E1. Photoactivation of AzMC led to a concentrationdependent loss in CYP2E1 activity and structural integrity resulting from the modification of both heme and protein. The photolabeling reaction degraded heme and produced a possible heme adduct. Probe incorporation into the protein occurred at multiple sites within substrate recognition sequence 5 (SRS-5). Based on a CYP2E1 homology model, we hypothesize AzMC labels SRS-5 residues, Leu363, Val364 and Leu368, in the active site. In addition, we propose a series of phenylalanines, especially Phe106, mediate contacts with the coumarin.

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Accumulation and drainage of hemin in the red cell membrane

Cited by 93 publications

References 27 publications

The Hog1 Mitogen-Activated Protein Kinase Mediates a Hypoxic Response inSaccharomyces cerevisiae

The Hog1 Mitogen-Activated Protein Kinase Mediates a Hypoxic Response inSaccharomyces cerevisiae

Clotrimazole Binds to Heme and Enhances Heme-dependent Hemolysis

CYP2E1 active site residues in substrate recognition sequence 5 identified by photoaffinity labeling and homology modeling

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