Encapsulation of concentrated hemoglobin solution in phospholipid vesicles retards the reaction with NO, but not CO, by intracellular diffusion barrier

Hiromi Sakai, Atsushi Sato, Kaoru Masuda, Shinji Takeoka, Eishun Tsuchida

    Research output: Contribution to journalArticle

    55 Citations (Scopus)

    Abstract

    One physiological significance of the red blood cell (RBC) structure is that NO binding of Hb is retarded by encapsulation with the cell membrane. To clarify the mechanism, we analyzed Hb-vesicles (HbVs) with different intracellular Hb concentrations, [Hb]in, and different particle sizes using stopped-flow spectrophotometry. The apparent NO binding rate constant, k′on (NO), of HbV at [Hb]in = 1 g/dl was 2.6 × 107 M-1 s-1, which was almost equal to kon (NO) of molecular Hb, indicating that the lipid membrane presents no obstacle for NO binding. With increasing [Hb]in to 35 g/dl, k′on (NO) decreased to 0.9 × 107 M-1 s-1, which was further decreased to 0.5 × 107 M-1 s-1 with enlarging particle diameter from 265 to 452 nm. For CO binding, which is intrinsically much slower than NO binding, k′on (CO) did not change greatly with [Hb]in and the particle diameter. Results obtained using diffusion simulations coupled with elementary binding reactions concur with these tendencies and clarify that NO is trapped rapidly by Hb from the interior surface region to the core of HbV at a high [Hb]in, retarding NO diffusion toward the core of HbV. In contrast, slow CO binding allows time for further CO-diffusion to the core. Simulations extrapolated to larger particles (8 μm) showing retardation even for CO binding. The obtained k′on (NO) and k′on (CO) yield values similar to those reported for RBCs. In summary, the intracellular, not extracellular, diffusion barrier is predominant due to the rapid NO binding that induces a rapid sink of NO from the interior surface to the core, retarding further NO diffusion and binding.

    Original languageEnglish
    Pages (from-to)1508-1517
    Number of pages10
    JournalJournal of Biological Chemistry
    Volume283
    Issue number3
    DOIs
    Publication statusPublished - 2008 Jan 18

    Fingerprint

    Diffusion barriers
    Carbon Monoxide
    Encapsulation
    Phospholipids
    Hemoglobins
    Spectrophotometry
    Cell membranes
    Membrane Lipids
    Particle Size
    Rate constants
    Blood
    Erythrocytes
    Particle size
    Cells
    Cell Membrane

    ASJC Scopus subject areas

    • Biochemistry

    Cite this

    Encapsulation of concentrated hemoglobin solution in phospholipid vesicles retards the reaction with NO, but not CO, by intracellular diffusion barrier. / Sakai, Hiromi; Sato, Atsushi; Masuda, Kaoru; Takeoka, Shinji; Tsuchida, Eishun.

    In: Journal of Biological Chemistry, Vol. 283, No. 3, 18.01.2008, p. 1508-1517.

    Research output: Contribution to journalArticle

    @article{1632fd68245b4e43a429cbbc596fe44c,
    title = "Encapsulation of concentrated hemoglobin solution in phospholipid vesicles retards the reaction with NO, but not CO, by intracellular diffusion barrier",
    abstract = "One physiological significance of the red blood cell (RBC) structure is that NO binding of Hb is retarded by encapsulation with the cell membrane. To clarify the mechanism, we analyzed Hb-vesicles (HbVs) with different intracellular Hb concentrations, [Hb]in, and different particle sizes using stopped-flow spectrophotometry. The apparent NO binding rate constant, k′on (NO), of HbV at [Hb]in = 1 g/dl was 2.6 × 107 M-1 s-1, which was almost equal to kon (NO) of molecular Hb, indicating that the lipid membrane presents no obstacle for NO binding. With increasing [Hb]in to 35 g/dl, k′on (NO) decreased to 0.9 × 107 M-1 s-1, which was further decreased to 0.5 × 107 M-1 s-1 with enlarging particle diameter from 265 to 452 nm. For CO binding, which is intrinsically much slower than NO binding, k′on (CO) did not change greatly with [Hb]in and the particle diameter. Results obtained using diffusion simulations coupled with elementary binding reactions concur with these tendencies and clarify that NO is trapped rapidly by Hb from the interior surface region to the core of HbV at a high [Hb]in, retarding NO diffusion toward the core of HbV. In contrast, slow CO binding allows time for further CO-diffusion to the core. Simulations extrapolated to larger particles (8 μm) showing retardation even for CO binding. The obtained k′on (NO) and k′on (CO) yield values similar to those reported for RBCs. In summary, the intracellular, not extracellular, diffusion barrier is predominant due to the rapid NO binding that induces a rapid sink of NO from the interior surface to the core, retarding further NO diffusion and binding.",
    author = "Hiromi Sakai and Atsushi Sato and Kaoru Masuda and Shinji Takeoka and Eishun Tsuchida",
    year = "2008",
    month = "1",
    day = "18",
    doi = "10.1074/jbc.M707660200",
    language = "English",
    volume = "283",
    pages = "1508--1517",
    journal = "Journal of Biological Chemistry",
    issn = "0021-9258",
    publisher = "American Society for Biochemistry and Molecular Biology Inc.",
    number = "3",

    }

    TY - JOUR

    T1 - Encapsulation of concentrated hemoglobin solution in phospholipid vesicles retards the reaction with NO, but not CO, by intracellular diffusion barrier

    AU - Sakai, Hiromi

    AU - Sato, Atsushi

    AU - Masuda, Kaoru

    AU - Takeoka, Shinji

    AU - Tsuchida, Eishun

    PY - 2008/1/18

    Y1 - 2008/1/18

    N2 - One physiological significance of the red blood cell (RBC) structure is that NO binding of Hb is retarded by encapsulation with the cell membrane. To clarify the mechanism, we analyzed Hb-vesicles (HbVs) with different intracellular Hb concentrations, [Hb]in, and different particle sizes using stopped-flow spectrophotometry. The apparent NO binding rate constant, k′on (NO), of HbV at [Hb]in = 1 g/dl was 2.6 × 107 M-1 s-1, which was almost equal to kon (NO) of molecular Hb, indicating that the lipid membrane presents no obstacle for NO binding. With increasing [Hb]in to 35 g/dl, k′on (NO) decreased to 0.9 × 107 M-1 s-1, which was further decreased to 0.5 × 107 M-1 s-1 with enlarging particle diameter from 265 to 452 nm. For CO binding, which is intrinsically much slower than NO binding, k′on (CO) did not change greatly with [Hb]in and the particle diameter. Results obtained using diffusion simulations coupled with elementary binding reactions concur with these tendencies and clarify that NO is trapped rapidly by Hb from the interior surface region to the core of HbV at a high [Hb]in, retarding NO diffusion toward the core of HbV. In contrast, slow CO binding allows time for further CO-diffusion to the core. Simulations extrapolated to larger particles (8 μm) showing retardation even for CO binding. The obtained k′on (NO) and k′on (CO) yield values similar to those reported for RBCs. In summary, the intracellular, not extracellular, diffusion barrier is predominant due to the rapid NO binding that induces a rapid sink of NO from the interior surface to the core, retarding further NO diffusion and binding.

    AB - One physiological significance of the red blood cell (RBC) structure is that NO binding of Hb is retarded by encapsulation with the cell membrane. To clarify the mechanism, we analyzed Hb-vesicles (HbVs) with different intracellular Hb concentrations, [Hb]in, and different particle sizes using stopped-flow spectrophotometry. The apparent NO binding rate constant, k′on (NO), of HbV at [Hb]in = 1 g/dl was 2.6 × 107 M-1 s-1, which was almost equal to kon (NO) of molecular Hb, indicating that the lipid membrane presents no obstacle for NO binding. With increasing [Hb]in to 35 g/dl, k′on (NO) decreased to 0.9 × 107 M-1 s-1, which was further decreased to 0.5 × 107 M-1 s-1 with enlarging particle diameter from 265 to 452 nm. For CO binding, which is intrinsically much slower than NO binding, k′on (CO) did not change greatly with [Hb]in and the particle diameter. Results obtained using diffusion simulations coupled with elementary binding reactions concur with these tendencies and clarify that NO is trapped rapidly by Hb from the interior surface region to the core of HbV at a high [Hb]in, retarding NO diffusion toward the core of HbV. In contrast, slow CO binding allows time for further CO-diffusion to the core. Simulations extrapolated to larger particles (8 μm) showing retardation even for CO binding. The obtained k′on (NO) and k′on (CO) yield values similar to those reported for RBCs. In summary, the intracellular, not extracellular, diffusion barrier is predominant due to the rapid NO binding that induces a rapid sink of NO from the interior surface to the core, retarding further NO diffusion and binding.

    UR - http://www.scopus.com/inward/record.url?scp=38349132524&partnerID=8YFLogxK

    UR - http://www.scopus.com/inward/citedby.url?scp=38349132524&partnerID=8YFLogxK

    U2 - 10.1074/jbc.M707660200

    DO - 10.1074/jbc.M707660200

    M3 - Article

    VL - 283

    SP - 1508

    EP - 1517

    JO - Journal of Biological Chemistry

    JF - Journal of Biological Chemistry

    SN - 0021-9258

    IS - 3

    ER -