Stabilization of the tetrameric structure of human and bovine hemoglobins by pseudocrosslinking with muconic acid

Anna Razynska, Barbara Matheson-Urbaitis, Clara Fronticelli, John H. Collins, Enrico Bucci

Research output: Contribution to journalArticlepeer-review

Abstract

In previous studies mono-3,5-dibromosalicyl-fumarate was used to introduce an intramolecular crosslink (pseudo-crosslink) in the β cleft between hemoglobin β subunits. Sedimentation velocity analysis indicated that the product had a mean molecular weight indicating a tetramer with low dissociability. The product had a P50 higher than that of native hemoglobin and a plasma retention time in the rat of about 3 h, i.e., four times longer than untreated hemoglobin. However, the product contained a fraction which was rapidly eliminated in the urine and which had a short plasma half-time of about 20 min, indicating the presence of a dissociable fraction. We have attempted to further enhance the tetrameric stability of hemoglobin and prevent urine elimination by positioning a longer chain carboxylic acid than fumaric acid into the β cleft. We reason that a longer molecule would allow for greater stabilizing interactions across the β cleft. In the present study human and bovine hemoglobins were reacted with mono-3-5-dibromosalicyl muconate. Muconic acid is two carbons longer than fumaric acid. The products were acylated at the β82 (human) and β81 (bovine) lysines of the β-cleft and had a low degree of dissociability. For reasons not presently understood, urine excretion was high and plasma half-time was not increased above that of untreated hemoglobin. In conclusion, it appears that only covalently crosslinked hemoglobins which are completely nondissociable tetramers escape filtration; tetramers with any degree of dissociability into dimers are filterable.

Original languageEnglish (US)
Pages (from-to)119-125
Number of pages7
JournalArchives of Biochemistry and Biophysics
Volume326
Issue number1
DOIs
StatePublished - Feb 1 1996
Externally publishedYes

Keywords

  • 3,5-dibromosalicylate
  • blood substitutes
  • hemoglobin dissociability
  • muconyl-hemoglobin
  • pseudocrosslinked hemoglobin
  • red cell substitutes
  • retention time

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology

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