scholarly journals Some effects of post-translational N-terminal acetylation of the human embryonic ζ globin protein

1995 ◽  
Vol 310 (2) ◽  
pp. 597-600 ◽  
Author(s):  
A Scheepens ◽  
R Mould ◽  
O Hofmann ◽  
T Brittain
Keyword(s):  
Oxygen Affinity ◽  
Chloride Ions ◽  
Site Directed Mutagenesis ◽  
Mutant Form ◽  
Oxygen Binding ◽  
Mutation Process ◽  
Zeta Chain ◽  
Major Disruption ◽  
Alkaline Bohr Effect

Using site-directed mutagenesis we have produced the first mutant form of a human embryonic haemoglobin. We have mutated the N-terminal Ser residue of the zeta-chain of haemoglobin Portland, zeta 2 gamma 2, (which is normally acetylated) to a Val (which possesses a free amine terminus). The protein spontaneously assembles into a fully functional tetramer which shows cooperative oxygen binding. Determination of the reactivity of the mutant protein with 2,3-diphosphoglycerate indicates that the mutation process does not lead to any major disruption of the protein structure. A comparison of the properties of the mutant and wild-type proteins identifies a significant role for the normal N-terminal acetylation of the zeta-chain with regard to the alkaline Bohr effect and the sensitivity of the oxygen affinity of the protein towards chloride ions. The possible physiological significance of this modification is discussed.

Studies of haemoglobin functions by site-directed mutagenesis

1986 ◽  
Vol 317 (1540) ◽  
pp. 443-447 ◽  
Keyword(s):  
Expression Vector ◽  
Oxygen Affinity ◽  
Coagulation Factor ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Oxygen Binding ◽  
Factor X ◽  
Binding Properties ◽  
High Oxygen Affinity ◽  
Alkaline Bohr Effect

Human (3-globin was synthesized in Escherichia coli as a cleavable fusion protein by using the expression vector pLcIIFX|3-globin(nic - ). The authentic (3-globin was liberated by digestion with blood coagulation factor X a and a 2 (3 2 tetramers were reconstituted. The oxygen-binding properties of reconstituted haemoglobin (Hb) were essentially the same as those of human native Hb. Two mutant haemoglobins were constructed by site-directed mutagenesis. HbNymphéas (Cys-93(3->Ser) showed a slightly increased oxygen affinity and diminished co-operativity with normal DPG (2,3-diphosphoglycerate) effect and slightly reduced alkaline Bohr effects. Hb Daphne (Cys-93(3->-Ser, His-143(3->- Arg) showed low co-operativity with high oxygen affinity. The alkaline Bohr effect was slightly reduced, but the DPG effect was enhanced by 50% by the His-143(3^ Arg mutation.

scholarly journals Hemoglobin Koln occurring in association with a beta zero thalassemia: hematologic and functional consequences

Blood ◽  
1989 ◽  
Vol 74 (1) ◽  
pp. 496-500
Author(s):  
F Galacteros ◽  
D Loukopoulos ◽  
P Fessas ◽  
J Kister ◽  
N Arous ◽  
...  
Keyword(s):  
Oxygen Affinity ◽  
Red Cells ◽  
Beta Thalassemia ◽  
Oxygen Binding ◽  
Morphological Alterations ◽  
Functional Consequences ◽  
High Oxygen Affinity ◽  
Alkaline Bohr Effect ◽  
Chain Synthesis ◽  
Globin Chain Synthesis

Hemoglobin (Hb) Koln-beta zero thalassemia compound heterozygosity was discovered in a young Greek patient. This gave us the unique opportunity for studying the functional properties of this unstable high-oxygen affinity hemoglobin variant in red cells containing almost pure Hb Koln. The red cells of the proposita exhibit morphological alterations and hematologic indices corresponding to the presence of an unstable Hb and beta thalassemia. Globin chain synthesis confirmed the association with a beta zero thalassemia gene. Oxygen-binding curves for these cells were biphasic, indicating the presence of both heme- saturated and of approximately 20% of non-cooperative Hb Koln. The major component exhibits an increased oxygen affinity, reduced cooperativeness, and normal alkaline Bohr effect. The 35-year-old proposita is active, has not been splenectomized, and has not been transfused in several years.

scholarly journals Hemoglobin Koln occurring in association with a beta zero thalassemia: hematologic and functional consequences

Blood ◽  
1989 ◽  
Vol 74 (1) ◽  
pp. 496-500 ◽  
Author(s):  
F Galacteros ◽  
D Loukopoulos ◽  
P Fessas ◽  
J Kister ◽  
N Arous ◽  
...  
Keyword(s):  
Oxygen Affinity ◽  
Red Cells ◽  
Beta Thalassemia ◽  
Oxygen Binding ◽  
Morphological Alterations ◽  
Functional Consequences ◽  
High Oxygen Affinity ◽  
Alkaline Bohr Effect ◽  
Chain Synthesis ◽  
Globin Chain Synthesis

Abstract Hemoglobin (Hb) Koln-beta zero thalassemia compound heterozygosity was discovered in a young Greek patient. This gave us the unique opportunity for studying the functional properties of this unstable high-oxygen affinity hemoglobin variant in red cells containing almost pure Hb Koln. The red cells of the proposita exhibit morphological alterations and hematologic indices corresponding to the presence of an unstable Hb and beta thalassemia. Globin chain synthesis confirmed the association with a beta zero thalassemia gene. Oxygen-binding curves for these cells were biphasic, indicating the presence of both heme- saturated and of approximately 20% of non-cooperative Hb Koln. The major component exhibits an increased oxygen affinity, reduced cooperativeness, and normal alkaline Bohr effect. The 35-year-old proposita is active, has not been splenectomized, and has not been transfused in several years.

Determination of interdependent ligand effects on human red cell oxygen affinity

1982 ◽  
Vol 42 (4) ◽  
pp. 339-345
Author(s):  
W. G. Zijlstra ◽  
B. Oeseburg ◽  
G. Kwant ◽  
A. Zwart
Keyword(s):  
Oxygen Affinity ◽  
Red Cell ◽  
Ligand Effects

Selectivity coefficients of some simple and complex inorganic ions for Spheron DEAE

1984 ◽  
Vol 49 (10) ◽  
pp. 2349-2354 ◽  
Author(s):  
František Vláčil ◽  
Karel Koňák
Keyword(s):  
Ion Exchange ◽  
Mobile Phase ◽  
Dynamic Method ◽  
Precious Metals ◽  
Inorganic Ions ◽  
Chromatographic Determination ◽  
Chloride Ions ◽  
Chloro Complexes ◽  
Selectivity Coefficients

The selectivity coefficients of the nitrate and chloride ions and of anionic chloro complexes of Au(III), Rh(III), Pd(II), and Pt(IV) for ion exchange on Spheron DEAE in the chloride form are determined by the dynamic method. the complex anion species formed are identified and the ion exchange nature of the sorption of precious metals on this sorbent is confirmed based on the elution order of the precious metals as determined previously by the column chromatography on Spheron DEAE using hydrochloric acid as the mobile phase. The effect of the presence of perchlorate in the mobile phase during the liquid chromatography of precious metals and during the chromatographic determination of nitrate traces is explained.

scholarly journals Crystallization and X-ray diffraction data analysis of human deoxyhaemoglobin A0 fully stripped of any anions

1999 ◽  
Vol 55 (11) ◽  
pp. 1914-1916 ◽  
Author(s):  
F. A. V. Seixas ◽  
W. F. de Azevedo ◽  
M. F. Colombo
Keyword(s):  
Diffraction Data ◽  
Oxygen Affinity ◽  
X Ray Diffraction ◽  
Structural Explanation ◽  
Human Haemoglobin ◽  
X Ray ◽  
High Resolution Structure ◽  
Allosteric Properties

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.

scholarly journals Determination of the Catalytic Base in Family 48 Glycosyl Hydrolases

2011 ◽  
Vol 77 (17) ◽  
pp. 6274-6276 ◽  
Author(s):  
Maxim Kostylev ◽  
David B. Wilson
Keyword(s):  
Aspartic Acid ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Thermobifida Fusca ◽  
Glycosyl Hydrolases ◽  
Content Type ◽  
Modeling Data ◽  
Partial Activity

ABSTRACTThe catalytic base in family 48 glycosyl hydrolases has not been previously established experimentally. Based on structural and modeling data published to date, we used site-directed mutagenesis and azide rescue activity assays to show definitively that the catalytic base inThermobifida fuscaCel48A is aspartic acid 225. Of the tested mutants, only Cel48A with the D225E mutation retained partial activity on soluble and insoluble substrates. In azide rescue experiments, only the D225G mutation, in the smallest residue tested, showed an increase in activity with added azide.

scholarly journals Haemoglobin polymorphisms affect the oxygen-binding properties in Atlantic cod populations

2008 ◽  
Vol 276 (1658) ◽  
pp. 833-841 ◽  
Author(s):  
Øivind Andersen ◽  
Ola Frang Wetten ◽  
Maria Cristina De Rosa ◽  
Carl Andre ◽  
Cristiana Carelli Alinovi ◽  
...  
Keyword(s):  
Evolutionary Biology ◽  
Quaternary Structure ◽  
Atlantic Cod ◽  
Three Dimensional ◽  
Oxygen Affinity ◽  
Oxygen Binding ◽  
Low Oxygen ◽  
Binding Properties ◽  
Important Species ◽  
The North

A major challenge in evolutionary biology is to identify the genes underlying adaptation. The oxygen-transporting haemoglobins directly link external conditions with metabolic needs and therefore represent a unique system for studying environmental effects on molecular evolution. We have discovered two haemoglobin polymorphisms in Atlantic cod populations inhabiting varying temperature and oxygen regimes in the North Atlantic. Three-dimensional modelling of the tetrameric haemoglobin structure demonstrated that the two amino acid replacements Met55β 1 Val and Lys62β 1 Ala are located at crucial positions of the α 1 β 1 subunit interface and haem pocket, respectively. The replacements are proposed to affect the oxygen-binding properties by modifying the haemoglobin quaternary structure and electrostatic feature. Intriguingly, the same molecular mechanism for facilitating oxygen binding is found in avian species adapted to high altitudes, illustrating convergent evolution in water- and air-breathing vertebrates to reduction in environmental oxygen availability. Cod populations inhabiting the cold Arctic waters and the low-oxygen Baltic Sea seem well adapted to these conditions by possessing the high oxygen affinity Val55–Ala62 haplotype, while the temperature-insensitive Met55–Lys62 haplotype predominates in the southern populations. The distinct distributions of the functionally different haemoglobin variants indicate that the present biogeography of this ecologically and economically important species might be seriously affected by global warming.

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