Carbodiimide-mediated coupling of benzenepentacarboxylate to human hemoglobin: structural and functional consequences
| Autor: | Joseph Bonaventura, Marius Brouwer, Robert Cashon |
|---|---|
| Rok vydání: | 1992 |
| Předmět: |
Chromatography
Molecular Structure Chemistry Covalent modification Hemoglobin A General Medicine Hydrogen-Ion Concentration Medicinal chemistry Dilution Coupling (electronics) Oxygen chemistry.chemical_compound Covalent bond Blood Substitutes Ethyldimethylaminopropyl Carbodiimide Benzene Derivatives Humans Hemoglobin Oxygen binding Carbodiimide |
| Zdroj: | Biomaterials, artificial cells, and immobilization biotechnology : official journal of the International Society for Artificial Cells and Immobilization Biotechnology. 20(2-4) |
| ISSN: | 1055-7172 |
| Popis: | We have examined the covalent modification of HbA with BPC (benzenepentacarboxylate) whose carboxyl groups were activated with EDC [1-ethyl-3-(3-dimethyl- aminopropyl)-carbodiimide]. Reaction of deoxy-HbA at pH 8 with a 10-fold excess of BPC, preactivated with a 2-fold excess of EDC for 5 minutes, followed by anion-exchange chromatography, gives three components with p50 values of 1.15 (unreacted HbA), 11.7 and 7.6 mm of Hg at 20 degrees C (50 mM Bis-Tris pH 7.0). Component III does not dissociate into dimers upon dilution, but components I and II do. When deoxy-HbA is reacted at pH 6 with 10-fold BPC, preactivated with two-fold EDC for 5 minutes, the resultant HbA derivatives can be separated into three components, with p50 values at pH 7 of 14.2, 10.2 and 5.2 mm of Hg, respectively. All three components are stable tetramers. Oxygen binding by all of the covalent HbA-(BPC)x complexes is cooperative, pH sensitive, but IHP insensitive. The latter observation suggest that BPC is covalently bound to HbA's DPG/IHP binding site. This conclusion is corroborated by reversed phase HPLC analysis which shows that all five modified HbAs contain at least one modified beta chain. In addition, 4 of the 5 derivatives also contain modified alpha chains. No inter or intratetramer crosslinks are observed. |
| Databáze: | OpenAIRE |
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