Molecular Weights of the Blood Pigments of Arenicola and of Lumbricus

Eriksson, Inga-Britta

doi:10.1038/130434a0

Cited by 11 publications

(4 citation statements)

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“…This would change our value of the molecular mass by less than 0.3%. Similarly, light-scattering measurements of Hb from the related earthworm (54), O. complanatum 1 , in the range 25-250 g/ml showed no concentration dependence.…”

Section: Molecular Weight Test Proteinsmentioning

confidence: 99%

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Assembly of the Gigantic Hemoglobin of the Earthworm Lumbricus terrestris

Zhu

Ownby

Riggs

et al. 1996

Journal of Biological Chemistry

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The extracellular hemoglobin of the earthworm Lumbricus terrestris has four major kinds of O 2 -binding chains: a, b, and c (forming a disulfide-linked trimer), and chain d. Non-heme, non-globin structural chains, "linkers," are also present. Light-scattering techniques have been used to show that the ferrous CO-saturated abc trimer and chain d form an (abcd) 4 complex of 285 kDa at neutral pH. Formation of the full-sized 4-MDa molecule requires the addition of linker chains in the proportion of two linkers per (abcd) 4 and occurs much more rapidly in the presence of 10 mM calcium. This stoichiometry is supported not only by direct quantitative analysis of the intact hemoglobin but also by the fact that the addition of 50% of the proposed stoichiometric quantity of linkers results in the conversion of 50% of the (abcd) 4 to full-sized molecules. Isolated COsaturated abc trimers self-associate to (abc) 2 and higher aggregates up to an apparent limit of (abc) 10 ϳ550 kDa. The extracellular hemoglobins of annelids were first shown by Svedberg and Eriksson (1) to be gigantic molecules of at least 3 MDa in molecular mass. Although several subsequent studies gave molecular masses close to 4 MDa or higher for the Hb of Lumbricus terrestris and related species (2-4), recent studies by scanning transmission electron microscopy (5) have suggested masses of 3.5-3.6 MDa, and early light-scattering measurements suggested even lower values (6, 7). A possible reason for this great variation in reported molecular masses is oxidation which has been shown to cause extensive dissociation of many annelid Hbs (8 -10). Goss et al. (9) used light scattering to show that oxidation of L. terrestris Hb at neutral pH caused a large drop in molecular weight. Similarly, Ascoli et al. (10) found that oxidation of the similar Hb from Octolasium complanatum 1 caused hemichrome formation and dissociation outside of a narrow range near pH 7.The stoichiometry has also been uncertain. The Hb of L. terrestris has four major kinds of O 2 -binding globin chains: a, b, and c (forming the disulfide-linked abc trimer) and chain d, together with non-heme structural chains, "linkers," designated L. Kapp et al. (11) and Vinogradov et al. (12) have reported that linkers comprise 33-36% of the total mass, but this conclusion was based largely on the staining of SDS-gels with Coomassie Blue. This dye, however, binds to different proteins to quite different extents (13,14). Ownby et al. (15) redetermined the stoichiometry by reverse-phase high performance liquid chromatography (HPLC).2 The weight proportions of linkers were found to be approximately 16.4% by two independent procedures: the integrated absorption of the HPLC peaks and amino acid analysis of these peaks. These results led to the conclusion that the overall stoichiometry is (abcd) 2 L. This stoichiometry is further supported by the results of SDS-capillary gel electrophoresis monitored at 214 nm (see our companion study (16)).The goals of the present experiments are to redetermine the molecu...

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Section: Molecular Weight Test Proteinsmentioning

confidence: 99%

“…Goss et al (9) used light scattering to show that oxidation of L. terrestris Hb at neutral pH caused a large drop in molecular weight. Similarly, Ascoli et al (10) found that oxidation of the similar Hb from Octolasium complanatum 1 caused hemichrome formation and dissociation outside of a narrow range near pH 7.…”

mentioning

confidence: 99%

Assembly of the Gigantic Hemoglobin of the Earthworm Lumbricus terrestris

Zhu

Ownby

Riggs

et al. 1996

Journal of Biological Chemistry

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“…In the case of O 2 , the presence of acellular Hb in the RBC-free layer, affects O 2 transport in two ways by augmenting the blood vessel wall partial pressure of O 2 (pO 2 ) gradient and oxygenated Hb (oxyHb) gradient, which therefore increases the net O 2 flux through the blood/tissue interface and which causes O 2 -dependent vasoconstriction [11–13]. Therefore inspired by our results with large sized polymeric Hbs that can overcome these limitations associated with the use of small sized cell-free Hbs, we have developed liposome/vesicle encapsulated Hb as an alternate strategy to provide a safe, efficacious and cost-effective alternative to acellular HBOCs [14]. …”

Section: Introductionmentioning

confidence: 99%

Hemoglobin Encapsulated Poly(Ethylene Glycol) Surface Conjugated Vesicles Attenuate Vasoactivity of Cell-Free Hemoglobin

Cabrales

Rameez²,

Palmer³

2012

CDDT

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Widespread clinical use of acellular hemoglobin (Hb)-based O2 carriers (HBOCs) has been hampered by their ability to elicit both vasoconstriction and systemic hypertension. This is primarily due to the ability of acellular Hb to extravasate through the blood vessel wall and scavenge endothelial-derived nitric oxide (NO). Encapsulation of Hb inside the aqueous core of liposomes retards the rates of NO dioxygenation and O2 release, which should reduce or eliminate the vasoactivity of Hb. Our aim is to determine the extent of systemic and microvascular vasoactive responses (hypertension, vasoconstriction and hypoperfusion) after infusion of vesicle encapsulated Hbs, in which the encapsulated Hb is in either the deoxygenated or carbon monoxide (CO) state (HbV and COHbV, respectively). To investigate this hypothesis, we used the hamster window chamber model subjected to two successive hypervolemic infusions of HbV and COHbV solutions (each infusion represents 10% of the animal’s calculated blood volume) at Hb concentrations of either 7 or 10 g/dL. The hypervolemic infusion model used in this study has all the regulatory mechanisms responsible for predicting the vasoconstrictive responses of HBOCs. The results of this study demonstrate the absence of vasoconstrictive and hypertensive responses upon single and multiple infusions of HbV and COHbV solutions. The HbV and COHbV solutions increased the plasma O2 carrying capacity. However, COHbV delivered low therapeutic levels of CO without inducing any microcirculatory disturbances. Significance: Vesicles containing Hb can be used as a new therapeutic agent in transfusion medicine to treat anemia and revert hypoperfusion.

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“…The classic sedimentation studies by Svedberg and Eriksson3 showed that annelid extracellular hemoglobins (Hbs), erythrocruorins, are gigantic molecules with molar masses of over 3 MDa. Levin4 and Roche5 first observed the remarkable two‐layered hexagonal images of the molecules by electron microscopy.…”

Section: Introductionmentioning

confidence: 99%

Linker chains of the gigantic hemoglobin of the earthworm Lumbricus terrestris: Primary structures of linkers L2, L3, and L4 and analysis of the connectivity of the disulfide bonds in linker L1

et al. 2006

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The extracellular hemoglobin (Hb) of the earthworm, Lumbricus terrestris, has four major kinds of globin chains: a, b, c, and d, present in equimolar proportions, and additional non-heme, non-globin scaffolding chains called linkers that are required for the calcium-dependent assembly of the full-sized molecule. The amino acid sequences of all four of the globin chains and one of the linkers (L1) have previously been determined. The amino acid sequences via cDNA of each of the three remaining linkers, L2, L3, and L4, have been determined so that the sequences of all constituent polypeptides of the hemoglobin are now known. Each linker has a highly conserved cysteine-rich segment of approximately 40 residues that is homologous with the seven ligand-binding repeats of the human low-density lipoprotein receptor (LDLR). Analysis of linker L1 shows that the connectivity of the three disulfide bonds is exactly the same as in the LDLR ligand-binding repeats. The presence of a calcium-binding site comprising one glutamyl and three aspartyl residues in both the LDLR repeats and in the linkers supports the suggestion that calcium is required for the folding and disulfide connectivity of the linkers as in the LDLR repeats. Linker L2 is markedly heterogeneous and contains unusual glycine-rich sequences near the NH2-terminus and a polar zipper-like sequence with imperfect repeats of Asp-Asp-His at the carboxyl terminus. Similar Asp-Asp-His repeats have been found in a protein homologous to superoxide dismutase in the hemolymph of certain mussels. These repeats may function as metal-binding sites.

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Molecular Weights of the Blood Pigments of Arenicola and of Lumbricus

Cited by 11 publications

References 1 publication

Assembly of the Gigantic Hemoglobin of the Earthworm Lumbricus terrestris

Assembly of the Gigantic Hemoglobin of the Earthworm Lumbricus terrestris

Hemoglobin Encapsulated Poly(Ethylene Glycol) Surface Conjugated Vesicles Attenuate Vasoactivity of Cell-Free Hemoglobin

Linker chains of the gigantic hemoglobin of the earthworm Lumbricus terrestris: Primary structures of linkers L2, L3, and L4 and analysis of the connectivity of the disulfide bonds in linker L1

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