A recombinant polymeric hemoglobin with conformational, functional, and physiological characteristics of an in vivo O2transporter

Authors:
  • TMToshiaki Mito
    Toshiaki Mito
  • STSarah J. Texel
    Sarah J. Texel
  • ABAndrea Bellelli
    Andrea Bellelli
  • MNMasaaki Nemoto
    Masaaki Nemoto
  • RTRichard J. Traystman
    Richard J. Traystman
  • RKRaymond C. Koehler
    Raymond C. Koehler
  • WBWilliam S. Brinigar
    William S. Brinigar
  • CFClara Fronticelli
    Clara Fronticelli
Publisher website
Google Scholar
Abstract
With the objective of developing a recombinant oxygen carrier suitable for therapeutic applications, we have employed an Escherichia coli expression system to synthesize in high-yield hemoglobin (Hb) Minotaur, containing α-human and β-bovine chains. Polymerization of Hb Minotaur through S-S intermolecular cross-linking was obtained by introducing a Cys at position β9 and substituting the naturally occurring Cys. This homogeneous polymer, Hb Polytaur, has a molecular mass of ∼500 kDa and was resistant toward reducing agents present in blood. In mice, the circulating half-time (3 h) was fivefold greater than adult human Hb (HbA). The half-time of autooxidation measured in blood (46 h) exceeded the circulating retention time. Hypervolemic exchange transfusion resulted in increased arterial blood pressure similar to that with albumin. The increase in pressure was less than that obtained by transfusion of cross-linked tetrameric Hb known to undergo renovascular extravasation. The nitric oxide reactivity of Hb Polytaur was similar to HbA, suggesting that the diminished pressor response to Hb Polytaur was probably related to diminished extravasation. Transfusion of 3% Hb Polytaur during focal cerebral ischemia reduced infarct volume by 22%. Therefore, site-specific Cys insertion on the Hb surface results in uniform size polymers that do not produce the large pressor response seen with tetrameric Hb. Polymerization maintains physiologically relevant oxygen and heme affinity, stability toward denaturation and oxidation, and effective oxygen delivery as indicated by reduced cerebral ischemic damage.

This publication has 49 references indexed in Scilit: