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Kinetics of reversible reductive carbonylation of heme in human cystathionine β-synthase.
Biochemistry. 2013 Jul 02; 52(26):4553-62.B

Abstract

Cystathionine β-synthase (CBS) catalyzes the condensation of homocysteine with serine or cysteine to form cystathionine and water or hydrogen sulfide (H2S), respectively. In addition to pyridoxal phosphate, human CBS has a heme cofactor with cysteine and histidine as ligands. While Fe(III)-CBS is inert to exogenous ligands, Fe(II)-CBS can be reversibly inhibited by carbon monoxide (CO) and reoxidized by O2 to yield superoxide radical. In this study, we have examined the kinetics of Fe(II)CO-CBS formation and reoxidation. Reduction of Fe(III)-CBS by dithionite showed a square root dependence on concentration, indicating that the reductant species was the sulfur dioxide radical anion (SO2(•-)) that exists in rapid equilibrium with S2O4(2-). Formation of Fe(II)CO-CBS from Fe(II)-CBS and 1 mM CO occurred with a rate constant of (3.1 ± 0.4) × 10(-3) s(-1) (pH 7.4, 25 °C). The reaction of Fe(III)-CBS with the reduced form of the flavoprotein methionine synthase reductase in the presence of CO and NADPH resulted in its reduction and carbonylation to form Fe(II)CO-CBS. Fe(II)-CBS was formed as an intermediate with a rate constant of (9.3 ± 2.5) × 10(2) M(-1) s(-1). Reoxidation of Fe(II)CO-CBS by O2 was multiphasic. The major phase showed a hyperbolic dependence on O2 concentration. Although H2S is a product of the CBS reaction and a potential heme ligand, we did not find evidence of an effect of exogenous H2S on activity or heme binding. Reversible reduction of CBS by a physiologically relevant oxidoreductase is consistent with a regulatory role for the heme and could constitute a mechanism for cross talk among the CO, H2S, and superoxide signaling pathways.

Authors+Show Affiliations

Laboratorio de Enzimología, Facultad de Ciencias, Facultad de Medicina, Universidad de la República, Montevideo, Uruguay.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

23790103

Citation

Carballal, Sebastián, et al. "Kinetics of Reversible Reductive Carbonylation of Heme in Human Cystathionine Β-synthase." Biochemistry, vol. 52, no. 26, 2013, pp. 4553-62.
Carballal S, Cuevasanta E, Marmisolle I, et al. Kinetics of reversible reductive carbonylation of heme in human cystathionine β-synthase. Biochemistry. 2013;52(26):4553-62.
Carballal, S., Cuevasanta, E., Marmisolle, I., Kabil, O., Gherasim, C., Ballou, D. P., Banerjee, R., & Alvarez, B. (2013). Kinetics of reversible reductive carbonylation of heme in human cystathionine β-synthase. Biochemistry, 52(26), 4553-62. https://doi.org/10.1021/bi4004556
Carballal S, et al. Kinetics of Reversible Reductive Carbonylation of Heme in Human Cystathionine Β-synthase. Biochemistry. 2013 Jul 2;52(26):4553-62. PubMed PMID: 23790103.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Kinetics of reversible reductive carbonylation of heme in human cystathionine β-synthase. AU - Carballal,Sebastián, AU - Cuevasanta,Ernesto, AU - Marmisolle,Inés, AU - Kabil,Omer, AU - Gherasim,Carmen, AU - Ballou,David P, AU - Banerjee,Ruma, AU - Alvarez,Beatriz, Y1 - 2013/06/21/ PY - 2013/6/25/entrez PY - 2013/6/25/pubmed PY - 2013/9/11/medline SP - 4553 EP - 62 JF - Biochemistry JO - Biochemistry VL - 52 IS - 26 N2 - Cystathionine β-synthase (CBS) catalyzes the condensation of homocysteine with serine or cysteine to form cystathionine and water or hydrogen sulfide (H2S), respectively. In addition to pyridoxal phosphate, human CBS has a heme cofactor with cysteine and histidine as ligands. While Fe(III)-CBS is inert to exogenous ligands, Fe(II)-CBS can be reversibly inhibited by carbon monoxide (CO) and reoxidized by O2 to yield superoxide radical. In this study, we have examined the kinetics of Fe(II)CO-CBS formation and reoxidation. Reduction of Fe(III)-CBS by dithionite showed a square root dependence on concentration, indicating that the reductant species was the sulfur dioxide radical anion (SO2(•-)) that exists in rapid equilibrium with S2O4(2-). Formation of Fe(II)CO-CBS from Fe(II)-CBS and 1 mM CO occurred with a rate constant of (3.1 ± 0.4) × 10(-3) s(-1) (pH 7.4, 25 °C). The reaction of Fe(III)-CBS with the reduced form of the flavoprotein methionine synthase reductase in the presence of CO and NADPH resulted in its reduction and carbonylation to form Fe(II)CO-CBS. Fe(II)-CBS was formed as an intermediate with a rate constant of (9.3 ± 2.5) × 10(2) M(-1) s(-1). Reoxidation of Fe(II)CO-CBS by O2 was multiphasic. The major phase showed a hyperbolic dependence on O2 concentration. Although H2S is a product of the CBS reaction and a potential heme ligand, we did not find evidence of an effect of exogenous H2S on activity or heme binding. Reversible reduction of CBS by a physiologically relevant oxidoreductase is consistent with a regulatory role for the heme and could constitute a mechanism for cross talk among the CO, H2S, and superoxide signaling pathways. SN - 1520-4995 UR - https://www.unboundmedicine.com/medline/citation/23790103/Kinetics_of_reversible_reductive_carbonylation_of_heme_in_human_cystathionine_β_synthase_ L2 - https://doi.org/10.1021/bi4004556 DB - PRIME DP - Unbound Medicine ER -