Combining the influence of two low O2 affinity-inducing chemical modifications of the central cavity of hemoglobin

Parimala Nacharaju, Joel M. Friedman, Muthuchidambaram Prabhakaran, Seetharama A. Acharya, Belur N. Manjula

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Abstract

HexaPEGylated hemoglobin (Hb), a non-hypertensive Hb, exhibits high O 2 affinity, which makes it difficult for it to deliver the desired levels of oxygen to tissues. The PEGylation of very low O2 affinity Hbs is now contemplated as the strategy to generate PEGylated Hbs with intermediate levels of O2 affinity. Toward this goal, a doubly modified Hb with very low O2 affinity has been generated. The amino terminal of the β-chain of HbA is modified by 2-hydroxy, 3-phospho propylation first to generate a low oxygen affinity Hb, HPPr-HbA. The oxygen affinity of this Hb is insensitive to DPG and IHP. Molecular modeling studies indicated potential interactions between the covalently linked phosphate group and Lys-82 of the trans β-chain. To further modulate the oxygen affinity of Hb, the αα-fumaryl cross-bridge has been introduced into HPPr-HbA in the mid central cavity. The doubly modified HbA (αα-fumaryl-HPPr- HbA) exhibits an O2 affinity lower than that of either of the singly modified Hbs, with a partial additivity of the two modifications. The geminate recombination and the visible resonance Raman spectra of the photoproduct of αα-fumaryl-HPPr-HbA also reflect a degree of additive influence of each of these modifications. The two modifications induced a synergistic influence on the chemical reactivity of Cys-93(β). It is suggested that the doubly modified Hb has accessed the low affinity T-state that is non-responsive to effectors. The doubly modified Hb is considered as a potential candidate for generating PEGylated Hbs with an O2 affinity comparable to that of erythrocytes for developing blood substitutes.

Original languageEnglish (US)
Pages (from-to)4554-4564
Number of pages11
JournalBiochemistry
Volume46
Issue number15
DOIs
Publication statusPublished - Apr 17 2007

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ASJC Scopus subject areas

  • Biochemistry

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