The role of an amino acid triad at the entrance of the heme pocket in human serum albumin for O2 and CO binding to iron protoporphyrin IX

Abstract

Complexation of iron(II) protoporphyrin IX (Fe²⁺PP) into a genetically engineered heme pocket on recombinant human serum albumin (rHSA) creates an artificial hemoprotein which can bind O₂ reversibly at room temperature. Here we highlight a crucial role of a basic amino acid triad the entrance of the heme pocket in rHSA (Arg-114, His-146, Lys-190) for O₂ and CO binding to the prosthetic Fe²⁺PP group. Replacing His-146 and/or Lys-190 with Arg resolved the structured heterogeneity of the possible two complexing modes of the porphyrin and afforded a single O₂ and CO binding affinity. Resonance Raman spectra show only one geometry of the axial His coordination to the central ferrous ion of the Fe²⁺PP.