Comparison of human haemoglobin and globin in denaturation

In this work, initial crystallographic studies of human haemoglobin (Hb) crystallized in isoionic and oxygen-free PEG solution are presented. Under these conditions, functional measurements of the O2-linked binding of water molecules and release of protons have evidenced that Hb assumes an unforeseen new allosteric conformation. The determination of the high-resolution structure of the crystal of human deoxy-Hb fully stripped of anions may provide a structural explanation for the role of anions in the allosteric properties of Hb and, particularly, for the influence of chloride on the Bohr effect, the mechanism by which Hb oxygen affinity is regulated by pH. X-ray diffraction data were collected to 1.87 Å resolution using a synchrotron-radiation source. Crystals belong to the space group P21212 and preliminary analysis revealed the presence of one tetramer in the asymmetric unit. The structure is currently being refined using maximum-likelihood protocols.

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Reductive Activation of Carbon Tetrachloride by Human Haemoglobin

Alternatives to Laboratory Animals ◽

10.1177/026119299302100110 ◽

1993 ◽

Vol 21 (1) ◽

pp. 57-64

Author(s):

Roberta Ferrara ◽

Michela Rezzadore ◽

Stefano Cazzaro ◽

Roberto Tolando ◽

Maurizio Manno

Keyword(s):

Carbon Monoxide ◽

Carbon Tetrachloride ◽

Anaerobic Incubation ◽

Reductive Activation ◽

Anaerobic Conditions ◽

Prosthetic Group ◽

Experimental Conditions ◽

Human Haemoglobin ◽

Reductive Metabolism

The reductive metabolism of carbon tetrachloride (CC14) by human haemoglobin (Hb) was observed in vitro by absolute absorption spectra recorded under anaerobic conditions. The following results were obtained: 1) a decrease of the 430nm peak typical of free reduced Hb (Hb2+); 2) the formation of a shoulder of absorbance, attributable to the production of a complex between Hb2+ and a metabolite of CC14 carbon monoxide (Hb-CO); and 3) the oxidation of some Hb2+ to methaemoglobin (Hb3+). The concentration of these three forms — Hb2+, Hb-CO and Hb3+ — during anaerobic incubation of Hb with CC14 was calculated algebraically from the absolute spectra. CO production was then calculated from the concentration of Hb-CO, using a suitable calibration curve. Interestingly, under identical experimental conditions, a substrate-dependent loss of Hb-derived haem, but not of Hb itself nor of haem-derived porphyrin fluorescence, was measured. Preliminary HPLC studies to clarify the discrepancy and, in particular, the role and fate of the haem group, showed two substrate-dependent modified haem products. The results indicate that human Hb is able to catalyse the reductive activation of CCl4, and suggest that, during the process, its prosthetic group haem may be modified by CC14 metabolites to products which maintain a tetrapyrrolic structure but are unable to react with pyridine.

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Age-dependent changes in volume and haemoglobin content of erythrocytes in the carp (Cyprinus carpio L.)

Journal of Experimental Biology ◽

10.1242/jeb.141.1.133 ◽

1989 ◽

Vol 141 (1) ◽

pp. 133-149 ◽

Cited By ~ 2

Author(s):

W. Speckner ◽

J. F. Schindler ◽

C. Albers

Keyword(s):

Gel Filtration ◽

Linear Increase ◽

Human Erythrocytes ◽

Main Peak ◽

Red Cell ◽

Human Haemoglobin ◽

Cell Fractions ◽

Haemoglobin Content

Carp erythrocytes were fractionated by angle-head centrifugation which yielded fractions with a linear increase in density. Haematological examinations revealed that the heavier red blood cells of carp had greater volumes (MCV), more haemoglobin (MCH) and higher haemoglobin concentrations (MCHC) than light ones. The same experiments with human red cell fractions yielded a decrease in MCV, constant MCH and an increase in MCHC. Haemoglobin content in individual erythrocytes was also determined by scanning stage absorbance cytophotometry to establish the frequency distribution of the cellular haemoglobin contents. In carp, the distribution was symmetrical with the means increasing with density. No such change with cell density was found in human erythrocytes. Both carp and human erythrocytes incorporated [2-14C]glycine in vitro. After gel filtration, radioactivity was detected in carp, but not in human, haemoglobin fractions. 14C was found in all three haemoglobin fractions, obtained by isoelectric focusing, and was present in the haem and in the globin. [2-14C]glycine-labelled erythrocytes were reinjected into chronically cannulated carp and followed in vivo for several months. With time, the main peak of scintillation counts shifted from red cell fractions of low to high density. This is considered as evidence that density and age of red cells in carp are positively correlated and that erythrocytes can synthesize haemoglobin while circulating in the peripheral blood.

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The development of an artificial stool usable for the surveillance of faecal haemoglobin testing

Annals of Clinical Biochemistry International Journal of Laboratory Medicine ◽

10.1177/0004563217732587 ◽

2017 ◽

Vol 55 (3) ◽

pp. 394-399

Author(s):

Akira Miike ◽

Zensuke Ogawa ◽

Ikunosuke Sakurabayashi

Keyword(s):

Haemoglobin Concentration ◽

Occult Blood ◽

Glycerol Concentration ◽

Faecal Occult Blood Testing ◽

Blood Testing ◽

Human Haemoglobin ◽

Standard Material ◽

Faecal Occult Blood ◽

Faecal Occult ◽

Haemoglobin Testing

Background Faecal occult blood testing is an important diagnostic tool for the detection of colorectal cancer. However, it has not been standardized due to the absence of suitable specimens for surveillance. Methods We developed a ready-to-use artificial stool made from rice flour. This new artificial stool homogeneously contains not only human haemoglobin A0 (HbA0) but also glycerol as an internal standard material. After the collection of the artificial stool into a buffer, the haemoglobin concentration in dispersed solution was measured using a method based on the peroxidase like activity of haemoglobin. The glycerol concentration was measured using a commercially available triglyceride measurement kit. Results With regard to the haemoglobin stability, the decrease in the level of human haemoglobin in the artificial stool was <2% when it was stored at −80℃ for four months, −20℃ for two weeks, and 5℃ for two days. The artificial stool was easily collected with the collecting tubes of a commercially available faecal haemoglobin test kit. The weight of the collected artificial stool could be calculated by measuring the concentration of glycerol in the extracting solution of the collected stool sample. The haemoglobin concentrations could be adjusted based on their collection weights. Conclusions The artificial stool has a paste-like consistency and contains both haemoglobin and glycerol homogeneously. Furthermore, the measured haemoglobin concentration could be determined based on the collected stool weight, which was directly related to the glycerol concentration. These features make it a useful material for the surveillance of faecal occult blood testing.

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The structure of haemoglobin. IX. A three-dimensional Fourier synthesis at 5.5 Å resolution: description of the structure

Proceedings of the Royal Society of London Series A - Mathematical and Physical Sciences ◽

10.1098/rspa.1962.0002 ◽

1962 ◽

Vol 265 (1321) ◽

pp. 161-187 ◽

Cited By ~ 70

Keyword(s):

Tertiary Structure ◽

Three Dimensional ◽

Sperm Whale ◽

Anomalous Scattering ◽

Fourier Synthesis ◽

Human Haemoglobin ◽

Contour Maps ◽

To Come ◽

Polypeptide Chains ◽

Sperm Whale Myoglobin

The electron density distribution in the unit cell is calculated at intervals of approximately 2Å and plotted in a series of sections parallel to (010). The contour maps show that haemoglobin consists of four subunits in a tetrahedral array. The subunits are identical in pairs in accordance with the twofold symmetry of the molecule. The two pairs are very similar in structure, and the members of each pair closely resemble the molecule of sperm-whale myoglobin. The four haem groups lie in separate pockets at the surface of the molecule. The positions of the iron atoms are confirmed by comparison of observed and calculated anomalous scattering effects, which also serve to determine the absolute configuration of the molecule. The four subunits found by X-ray analysis correspond to the four polypeptide chains into which haemoglobin can be divided by chemical methods. In horse haemoglobin the amino acid sequence within these chains is still partly unknown, but in human haemoglobin it has already been determined. Comparison of this sequence with the tertiary structure of the chains as now revealed in horse haemoglobin and with the atomic model of sperm-whale myoglobin recently obtained by Kendrew and his collaborators shows many interesting relations. Prolines appear to come where the chains turn corners or where their configuration is known to be non-helical. On the other hand, the chains also have corners which contain no proline. Certain residues appear to be structurally vital, because they appear in identical positions in myoglobin and in the two chains of haemoglobin, while in other parts of the molecule a wide variety of different side-chains appears to be allowed.

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Mercury binding to human haemoglobin

Cellular and Molecular Life Sciences ◽

10.1007/bf01946793 ◽

1973 ◽

Vol 29 (9) ◽

pp. 1178-1179 ◽

Cited By ~ 9

Author(s):

D. Barltrop ◽

A. M. Smith

Keyword(s):

Human Haemoglobin

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