α‐Chain and Globin: Intermediates in the Synthesis of Rabbit Hemoglobin

Baglioni, Corrado; Campana, Theresa

doi:10.1111/j.1432-1033.1967.tb00162.x

Cited by 75 publications

(17 citation statements)

References 18 publications

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“…Given that a small pool of free a chains is known to exist in erythroblasts (8,20,40,64,77,93,97), Yu et al (119) hypothesized that the ill effects caused by Ahsp gene disruption might be mitigated by lowering a globin gene dosage. This would reduce or eliminate the pool of free a chains that are unescorted by AHSP, and consequently might lessen the severity of the Ahsp knockout phenotype (119).…”

Section: Role Of Ahsp As a Chaperone During Hemoglobin Productionmentioning

confidence: 99%

The Role of Alpha-Hemoglobin Stabilizing Protein in Redox Chemistry, Denaturation, and Hemoglobin Assembly

Mollan

Weiss

et al. 2010

Antioxidants & Redox Signaling

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Hemoglobin biosynthesis in erythrocyte precursors involves several steps. The correct ratios and concentrations of normal alpha (a) and beta (b) globin proteins must be expressed; apoproteins must be folded correctly; heme must be synthesized and incorporated into these globins rapidly; and the individual a and b subunits must be rapidly and correctly assembled into heterotetramers. These events occur on a large scale in vivo, and dysregulation causes serious clinical disorders such as thalassemia syndromes. Recent work has implicated a conserved erythroid protein known as Alpha-Hemoglobin Stabilizing Protein (AHSP) as a participant in these events. Current evidence suggests that AHSP enhances a subunit stability and diminishes its participation in harmful redox chemistry. There is also evidence that AHSP facilitates one or more early-stage post-translational hemoglobin biosynthetic events. In this review, recent experimental results are discussed in light of several current models describing globin subunit folding, heme uptake, assembly, and denaturation during hemoglobin synthesis. Particular attention is devoted to molecular interactions with AHSP that relate to a chain oxidation and the ability of a chains to associate with partner b chains. Antioxid. Redox Signal. 12, 219-232.

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Section: Role Of Ahsp As a Chaperone During Hemoglobin Productionmentioning

confidence: 99%

The Role of Alpha-Hemoglobin Stabilizing Protein in Redox Chemistry, Denaturation, and Hemoglobin Assembly

Mollan

Weiss

et al. 2010

Antioxidants & Redox Signaling

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“…Rabbit reticulocytes synthesize essentially equal amounts of the a and chains of hemoglobin (20)(21)(22). The experiments shown in Figs.…”

Section: Discussionmentioning

confidence: 91%

Synthesis of α and β Chains of Rabbit Hemoglobin in a Cell-Free Extract from Krebs II Ascites Cells

Housman

Pemberton

Taber

1971

Proc. Natl. Acad. Sci. U.S.A.

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An RNA sedimenting at 10 S with a molecular weight of 2.3 X 106, was isolated from rabbit reticylocyte polyribosomes. When this RNA is added to a cell-free extract from Krebs II ascites cells, both a and ,B chains of rabbit hemoglobin are synthesized; g chains are made in about 50% excess over a chains.Isolation and characterization of the messenger RNA for a and # chains of rabbit hemoglobin has been reported by several groups (1-7). The informational content of this RNA has been assayed by adding it to a eukaryotic cell-free extract that was already synthesizing a homologous hemoglobin (3, 7) or myosin (5), or to a bacterial cell-free extract (4). The protein made in these extracts was analyzed by separation of the polypeptide chains into those whose synthesis was directed by endogenous messenger RNA and those whose synthesis was the result of added RNA. We report here the synthesis of both a and 13 chains of rabbit hemoglobin when 10S messenger RNA from rabbit reticulocytes is added to a cell-free extract from Krebs II ascites cells. This cell-free extract can be incubated to eliminate the synthesis of endogenous protein, while it retains the ability to respond to added viral RNA (8,9). Our results extend the use of previously incubated extracts of Krebs II ascites cells to characterization of proteins encoded by isolated mammalian messenger RNAs. METHODSThe 10S messenger RNA was prepared from rabbit reticulocytes by the method of Labrie (2). Polyribosomes were dissociated in EDTA, and the ribonucleoprotein particle that contained 10S messenger RNA was separated from ribosomal subunits on a sucrose density gradient (2). The RNA was then precipitated by adjusting the appropriate fractions to 0.1 M NaCl and 0.5% sodium dodecyl sulfate (SDS), followed by the addition of 3 volumes of ethanol. After 2 hr at -20'C, the precipitate was collected by centrifugation, dissolved in 0.1 M NaCl-0.5% SDS-1 mM EDTA-0.01 M Tris [pH 7.4 (SDS buffer)], layered over a 15-30% sucrose density gradient made in the same buffer, and centrifuged for 21 hr at 22°C and 27,000 rpm in the Beckman SW 27 rotor. The gradient was monitored for Aseonm; fractions containing RNA sedimenting at 10 S were pooled and precipitated with ethanol.The precipitated RNA was dissolved in SDS buffer and extracted with phenol-chloroform, as described by Penman (10). After precipitation with ethanol, the RNA was washed at least three times with ethanol at -20°C to remove SDS, dissolved in water at a concentration of 1 mg/ml, and stored 2716 at -70°C. 40 ug of purified 10S RNA was prepared from 10 mg of polyribosomes. Maintenance of Krebs II ascites cells in mice has been described (8). A cell-free extract was prepared from these cells according to a method communicated to us by M. B. Matthews, and used by him and others (8, 9). Mice bearing tumors were killed 7-10 days after inoculation, and the ascites cells were removed from the peritoneal cavity into 0.146 M NaCl-35 mM Tris (pH 7.5). They were collected by centrifugation at 800 rpm (120 X g) for 2 ...

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“…It has been suggested that achains assist in the removal of completed f8-chains from polyribosomes (22,23 …”

Section: Methodsmentioning

confidence: 99%

“…Table I lists the patients whose blood was used for the study of P-globin m-RNA translation. Five patients of Italian descent, ages [11][12][13][14][15][16][17][18][19][20][21][22][23][24][25] yr, with Cooley's anemia or P-thalassemia major were examined. All had severe anemia and, except for one patient, required blood transfusions at 1-2 month intervals.…”

Section: Methodsmentioning

confidence: 99%

Translation of β-Globin m-RNA in β-Thalassemia and the S and C Hemoglobinopathies

Rieder¹

1972

J. Clin. Invest.

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A B S T R A C T Genetic and biochemical evidence indicates that in 13-thalassemia there is impaired synthesis of the P-globin chains of hemoglobin A. In patients heterozygous for the hemoglobinopathies, hemoglobin S and hemoglobin C, the mutant 1-chain is produced in smaller amounts than normal PI. Defective m-RNA translation has been suggested as a possible cause of decreased 1-globin polypeptide synthesis in thalassemia and the hemoglobinopathies. In the present study, the ribosomal assembly of 13-globin chains was examined in the peripheral, nucleated red blood cells and reticulocytes of patients with Cooley's anemia, thalassemia intermedia, sickle thalassemia, sickle cell anemia, hemoglobin C disease, and in hemolytic anemias not associated with a hemoglobinopathy. The translation times of PA Ps , and P' did not differ significantly (average times: pA = 75 sec. range 43-114, ps = 69 sec, pc = 92 sec).In thalassemia, no evidence was found for a delay in translation as the cause of the marked impairment of 13-globin synthesis. In several specimens of peripheral blood from thalassemic patients, the translation time of the P-chain was even shorter than in nonthalassemic specimens (average time = 45 sec, range 35-59). The results suggest that the defect in 13-globin synthesis in P-thalassemia is due to impaired initiation of 13-globin chain assembly or a quantitative deficiency in m-RNA.

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α‐Chain and Globin: Intermediates in the Synthesis of Rabbit Hemoglobin

Cited by 75 publications

References 18 publications

The Role of Alpha-Hemoglobin Stabilizing Protein in Redox Chemistry, Denaturation, and Hemoglobin Assembly

The Role of Alpha-Hemoglobin Stabilizing Protein in Redox Chemistry, Denaturation, and Hemoglobin Assembly

Synthesis of α and β Chains of Rabbit Hemoglobin in a Cell-Free Extract from Krebs II Ascites Cells

Translation of β-Globin m-RNA in β-Thalassemia and the S and C Hemoglobinopathies

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