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Allosteric transition intermediates modelled by crosslinked haemoglobins

M A Schumacher1, M M Dixon, R Kluger

  • 1Department of Biochemistry and Molecular Biology, Oregon Health Sciences University, Portland 97201-3098, USA.

Nature
|May 4, 1995
PubMed
Summary

Chemical crosslinking stabilizes short-lived hemoglobin intermediates. This study reveals the structures of these transitional states, offering a snapshot of the nascent high-oxygen-affinity (R) state in hemoglobin.

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Area of Science:

  • Biochemistry
  • Structural Biology
  • Biophysics

Background:

  • The transition between low-oxygen-affinity (T) and high-oxygen-affinity (R) states of hemoglobin is crucial for oxygen transport.
  • X-ray crystallography has defined the T and R states, but transient intermediates remain structurally elusive.

Purpose of the Study:

  • To structurally characterize short-lived hemoglobin intermediates using chemical crosslinking.
  • To capture and analyze the structural features of hemoglobin during its transition from the T to the R state.

Main Methods:

  • Chemical crosslinking of deoxyhemoglobin using 3,3'-stilbenedicarboxylic acid (S) or trimesic acid (Tm) between beta Val1 and beta Lys82 residues.
  • X-ray crystallography of crosslinked hemoglobin variants (alpha 2 beta 1S82 beta, alpha 2 beta 1Tm82 beta, and alpha 2 beta 1,82Tm82 beta) after carbon monoxide saturation.

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Main Results:

  • The alpha 2 beta 1S82 beta variant, crosslinked with S, adopted a complete R-state conformation with near-normal oxygen affinity.
  • The alpha 2 beta 1Tm82 beta and alpha 2 beta 1,82Tm82 beta variants, crosslinked with Tm, exhibited low oxygen affinity and structural features of transitional intermediates, indicating incomplete R-state transition.
  • These Tm-crosslinked variants represent a structural snapshot of the early R state.

Conclusions:

  • Chemical crosslinking is a viable method to stabilize and structurally characterize transient hemoglobin intermediates.
  • The study provides novel structural insights into the nascent R state of hemoglobin, elucidating key features of the transition mechanism.
  • Understanding these intermediates enhances our knowledge of allosteric regulation in hemoglobin and related proteins.