A cis-proline in alpha-hemoglobin stabilizing protein directs the structural reorganization of alpha-hemoglobin

J Biol Chem. 2009 Oct 23;284(43):29462-9. doi: 10.1074/jbc.M109.027045. Epub 2009 Aug 25.


alpha-Hemoglobin (alphaHb) stabilizing protein (AHSP) is expressed in erythropoietic tissues as an accessory factor in hemoglobin synthesis. AHSP forms a specific complex with alphaHb and suppresses the heme-catalyzed evolution of reactive oxygen species by converting alphaHb to a conformation in which the heme is coordinated at both axial positions by histidine side chains (bis-histidyl coordination). Currently, the detailed mechanism by which AHSP induces structural changes in alphaHb has not been determined. Here, we present x-ray crystallography, NMR spectroscopy, and mutagenesis data that identify, for the first time, the importance of an evolutionarily conserved proline, Pro(30), in loop 1 of AHSP. Mutation of Pro(30) to a variety of residue types results in reduced ability to convert alphaHb. In complex with alphaHb, AHSP Pro(30) adopts a cis-peptidyl conformation and makes contact with the N terminus of helix G in alphaHb. Mutations that stabilize the cis-peptidyl conformation of free AHSP, also enhance the alphaHb conversion activity. These findings suggest that AHSP loop 1 can transmit structural changes to the heme pocket of alphaHb, and, more generally, highlight the importance of cis-peptidyl prolyl residues in defining the conformation of regulatory protein loops.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Blood Proteins / chemistry*
  • Blood Proteins / genetics
  • Blood Proteins / metabolism
  • Crystallography, X-Ray
  • Hemoglobin A / chemistry*
  • Hemoglobin A / genetics
  • Hemoglobin A / metabolism
  • Humans
  • Molecular Chaperones / chemistry*
  • Molecular Chaperones / genetics
  • Molecular Chaperones / metabolism
  • Mutation
  • Nuclear Magnetic Resonance, Biomolecular
  • Proline / chemistry
  • Proline / genetics
  • Proline / metabolism
  • Protein Stability
  • Protein Structure, Quaternary / physiology
  • Protein Structure, Secondary / physiology
  • Structure-Activity Relationship


  • AHSP protein, human
  • Blood Proteins
  • Molecular Chaperones
  • Hemoglobin A
  • Proline

Associated data

  • PDB/3IA3