Haemoglobin binding with haptoglobin. Unequivocal demonstration that the beta-chains of human haemoglobin bind to haptoglobin.
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ABSTRACT:
SUBMITTER: Kazim AL
PROVIDER: S-EPMC1161299 | biostudies-other | 1980 Jan
REPOSITORIES: biostudies-other
Similar Datasets
Project description:A synthetic approach was employed to identify the haptoglobin-binding site on the alpha-chain of human haemoglobin. This approach cosists of the synthesis of a series of consecutive overlapping peptides that, together, systematically represent the entire protein chain. Fourteen peptides were synthesized (alpha 1-15, alpha 11-25, alpha 21-35, alpha 31-45, alpha 41-55, alpha 51-65, alpha 61-75, alpha 71-85, alpha 81-95, alpha 91-105, alpha 101-115, alpha 111-125, alpha 121-135 and alpha 131-141), and their ability bind human haptoglobin was studied, Only peptide alpha 121-135 bound haptoglobin significantly. On this basis we conclude that the haptoglobin-binding site on the alpha-chain of haemoglobin resides within, but does not necessarily encompass all of, the region alpha 121-135.
| S-EPMC1163152 | biostudies-other
Project description:A synthetic approach was employed to identify the haptoglobin-binding sites on the beta-chain of human haemoglobin. This approach consists of the synthesis of a series of consecutive overlapping peptides that, together, systematically represent the entire protein chain. Fourteen synthetic peptides (beta 1-15, beta 11-25 etc.) were examined for their ability to bind human haptoglobin by quantitative solid-phase radiometric titrations of 125I-labelled haptoglobin. Of these 14 peptides only peptides beta 11-25 and beta 131-146 bound haptoglobin significantly; peptide beta 21-35 exhibited a small binding activity as a consequence of the overlap with peptide beta 11-25. On this basis and by examination of the three-dimensional structure of haemoglobin, it was concluded that the beta-chain of haemoglobin has two binding sites for haptoglobin that reside in, but do not necessarily encompass all of, the regions beta 11-25 and beta 131-146.
| S-EPMC1146586 | biostudies-other
Project description:One of the difficulties in creating a blood substitute on the basis of human haemoglobin (Hb) is the toxic nature of Hb when it is outside the safe environment of the red blood cells. The plasma protein haptoglobin (Hp) takes care of the Hb physiologically leaked into the plasma - it binds Hb and makes it much less toxic while retaining the Hb's high oxygen transporting capacity. We used Electron Paramagnetic Resonance (EPR) spectroscopy to show that the protein bound radical induced by H 2O 2 in Hb and Hp-Hb complex is formed on the same tyrosine residue(s), but, in the complex, the radical is found in a more hydrophobic environment and decays slower than in unbound Hb, thus mitigating its oxidative capacity. The data obtained in this study might set new directions in engineering blood substitutes for transfusion that would have the oxygen transporting efficiency typical of Hb, but which would be non-toxic.
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Project description:The haptoglobin-haemoglobin receptor of the African trypanosome species, Trypanosoma brucei, is expressed when the parasite is in the bloodstream of the mammalian host, allowing it to acquire haem through the uptake of haptoglobin-haemoglobin complexes. Here we show that in Trypanosoma congolense this receptor is instead expressed in the epimastigote developmental stage that occurs in the tsetse fly, where it acts as a haemoglobin receptor. We also present the structure of the T. congolense receptor in complex with haemoglobin. This allows us to propose an evolutionary history for this receptor, charting the structural and cellular changes that took place as it adapted from a role in the insect to a new role in the mammalian host.
| S-EPMC4889325 | biostudies-literature
Project description:The adverse effects of extra-erythrocytic hemoglobin (Hb) are counterbalanced by several plasma proteins devoted to facilitate the clearance of free heme and Hb. In particular, haptoglobin (Hp) traps the αβ dimers of Hb, which are delivered to the reticulo-endothelial system by CD163 receptor-mediated endocytosis. Since Hp:Hb complexes show heme-based reactivity, kinetics of O2 dissociation from the ferrous oxygenated human Hp1-1:Hb and Hp2-2:Hb complexes (Hp1-1:Hb(II)-O2 and Hp2-2:Hb(II)-O2, respectively) have been determined. O2 dissociation from Hp1-1:Hb(II)-O2 and Hp2-2:Hb(III)-O2 follows a biphasic process. The relative amplitude of the fast and slow phases ranges between 0.47 and 0.53 of the total amplitude, with values of koff1 (ranging between 25.6 ± 1.4 s-1 and 29.1 ± 1.3 s-1) being about twice faster than those of koff2 (ranging between 13.8 ± 1.6 s-1 and 16.1 ± 1.2 s-1). Values of koff1 and koff2 are essentially the same independently on whether O2 dissociation has been followed after addition of a dithionite solution or after O2 displacement by a CO solution in the presence of dithionite. They correspond to those reported for the dissociation of the first O2 molecule from tetrameric Hb(II)-O2, indicating that in the R-state α and β chains are functionally heterogeneous and the tetramer and the dimer behave identically. Accordingly, the structural conformation of the α and β chains of the Hb dimer bound to Hp corresponds to that of the subunits of the Hb tetramer in the R-state.
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Project description:Haptoglobin binds haemoglobin so firmly that there is practically no dissociation. It would be expected that the heat of the reaction would be relatively large. The development of the microcalorimeter by Benzinger offered the opportunity to measure the heat of reaction. The experiments were carried out in the Beckman 190B Microcalorimeter in two ways: (1) a constant amount of haptoglobin (Kabi; 65mg.) with different amounts of haemoglobin, and (2) a constant amount of haemoglobin (32.5mg.) with different amounts of haptoglobin. The proteins, each in 5ml. of 0.15m-phosphate buffer, pH7.4, were placed in equal-volume calorimeter cells. The heat produced/mg. of haemoglobin was calculated from the slope of the curve for a constant amount of haptoglobin and from the maximum heat for a constant amount of haemoglobin. This heat is about 70kcal./mole at 37 degrees . DeltaH varies with temperature, being -70.2 at 37 degrees , -29.7 at 20 degrees and 7.2 at 4 degrees . From the amount of haptoglobin required to attain maximum heat with 32.5mg. of haemoglobin and the amount of haemoglobin required to attain maximum heat with 65mg. of haptoglobin, it appears that at excess of haptoglobin there is competition between the reactions of 2moles of haptoglobin with 1mole of haemoglobin (or 2 alphabeta-chains) and 1mole of haptoglobin with 1mole of haemoglobin.
| S-EPMC1185122 | biostudies-other
Project description:Haptoglobin is a plasma protein of mammals that plays a crucial role in vascular homeostasis by binding free haemoglobin released from ruptured red blood cells. Trypanosoma brucei can exploit this by internalising haptoglobin-haemoglobin complex to acquire host haem. Here, we investigated the impact of haptoglobin deficiency (Hp-/-) on T. brucei brucei infection and the parasite´s capacity to internalise haemoglobin in a Hp-/- mouse model. The infected Hp-/- mice exhibited normal disease progression, with minimal weight loss and no apparent organ pathology, similarly to control mice. While the proteomic profile of mouse sera significantly changed in response to T. b. brucei, no differences in the infection response markers of blood plasma between Hp-/- and control Black mice were observed. Similarly, very few quantitative differences were observed between the proteomes of parasites harvested from Hp-/- and Black mice, including both endogenous proteins and internalised host proteins. While haptoglobin was indeed absent from parasites isolated from Hp-/-mice, haemoglobin peptides were unexpectedly detected in parasites from both Hp-/- and Black mice. Combined, the data support the dispensability of haptoglobin for haemoglobin internalisation by T. b. brucei during infection in mice. Since the trypanosomes knock-outs for their haptoglobin-haemoglobin receptor (HpHbR) internalised significantly less haemoglobin from Hp-/- mice compared to those isolated from Black mice, it suggests that T. b. brucei employs also an HpHbR-independent haptoglobin-mediated mode for haemoglobin internalisation. Our study reveals a so-far hidden flexibility of haemoglobin acquisition by T. b. brucei and offers novel insights into alternative haemoglobin uptake pathways.
| S-EPMC11304504 | biostudies-literature
Project description:Trypanosoma brucei requires haemoglobin for survival in the mammalian bloodstream. Haptoglobin (Hp), a host protein known to bind haemoglobin, was determined as a key component for haemoglobin acquisition by the parasite. Here, we investigated the impact of haptoglobin deficiency on T. brucei brucei infection and the parasite´s capacity to internalise haemoglobin in the Hp-/- mouse model. The infected Hp-/- mice exhibited normal disease progression, with minimal weight loss and no apparent organ pathology similarly to control mice. While the proteomic profile of mouse sera significantly changed in response to T. b. brucei, no differences in the infection response markers of blood plasma have been observed between Hp-/- and control Black mice. Similarly, very few quantitative differences have been observed between the proteomes of parasites harvested from Hp-/- and Black mice, entailing both endogenous proteins and internalised host proteins. While haptoglobin was indeed absent from parasites isolated from Hp-/-mice, haemoglobin peptides were unexpectedly detected in parasites from both Hp-/- and Black mice. This was verified by western blotting, EM immunolabeling of parasites cryosections, and analytical quantification of haem cofactors in the parasites extracts. Combined, the data support the Hp dispensability for haemoglobin internalisation by T. b. brucei during infection in mice. To further explore this phenomenon, we have generated T. b. brucei haptoglobin-haemoglobin receptor (HpHbR) knock-outs and used them to infect Hp-/- and control mice. The HpHbR-knock-out trypanosomes internalised significantly less haemoglobin from Hp-/- mice compared to those isolated from Black mice strongly suggesting that T. b. brucei employs an HpHbR-independent Hp-mediated mode for haemoglobin internalisation. Our study reveals a so-far hidden flexibility of haemoglobin acquisition by T. b. brucei and offers novel insights into alternative haemoglobin uptake pathways.
2024-07-16 | PXD052778 | Pride
Project description:Non-typeable Haemophilus influenzae (NTHi) is a major human pathogen for which there is no globally licensed vaccine. NTHi has a strict growth requirement for iron and encodes several systems to scavenge elemental iron and heme from the host. An effective NTHi vaccine would target conserved, essential surface factors, such as those involved in iron acquisition. Haemoglobin-haptoglobin binding proteins (Hgps) are iron-uptake proteins localized on the outer-membrane of NTHi. If the Hgps are to be included as components of a rationally designed subunit vaccine against NTHi, it is important to understand their prevalence and diversity. Following analysis of all available Hgp sequences, we propose a standardized grouping method for Hgps, and demonstrate increased diversity of these proteins than previously determined. This analysis demonstrated that genes encoding variants HgpB and HgpC are present in all strains examined, and almost 40% of strains had a duplicate, nonidentical hgpB gene. Hgps are also phase-variably expressed; the encoding genes contain a CCAA(n) simple DNA sequence repeat tract, resulting in biphasic ON-OFF switching of expression. Examination of the ON-OFF state of hgpB and hgpC genes in a collection of invasive NTHi isolates demonstrated that 58% of isolates had at least one of hgpB or hgpC expressed (ON). Varying expression of a diverse repertoire of hgp genes would provide strains a method of evading an immune response while maintaining the ability to acquire iron via heme. Structural analysis of Hgps also revealed high sequence variability at the sites predicted to be surface exposed, demonstrating a further mechanism to evade the immune system-through varying the surface, immune-exposed regions of the membrane anchored protein. This information will direct and inform the choice of candidates to include in a vaccine against NTHi.
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Project description:After subarachnoid haemorrhage, prolonged exposure to toxic extracellular haemoglobin occurs in the brain. Here, we investigate the role of haemoglobin neurotoxicity in vivo and its prevention. In humans after subarachnoid haemorrhage, haemoglobin in cerebrospinal fluid was associated with neurofilament light chain, a marker of neuronal damage. Most haemoglobin was not complexed with haptoglobin, an endogenous haemoglobin scavenger present at very low concentration in the brain. Exogenously added haptoglobin bound most uncomplexed haemoglobin, in the first 2 weeks after human subarachnoid haemorrhage, indicating a wide therapeutic window. In mice, the behavioural, vascular, cellular and molecular changes seen after human subarachnoid haemorrhage were recapitulated by modelling a single aspect of subarachnoid haemorrhage: prolonged intrathecal exposure to haemoglobin. Haemoglobin-induced behavioural deficits and astrocytic, microglial and synaptic changes were attenuated by haptoglobin. Haptoglobin treatment did not attenuate large-vessel vasospasm, yet improved clinical outcome by restricting diffusion of haemoglobin into the parenchyma and reducing small-vessel vasospasm. In summary, haemoglobin toxicity is of clinical importance and preventable by haptoglobin, independent of large-vessel vasospasm.
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