Analysis of the contribution of the globin and reductase domains to the ligand-binding properties of bacterial haemoglobins.
Bacterial Hbs (haemoglobins), like VHb (Vitreoscilla sp. Hb), and flavoHbs (flavohaemoglobins), such as FHP (Ralstonia eutropha flavoHb), have different autoxidation and ligand-binding rates. To determine the influence of each domain of flavoHbs on ligand binding, we have studied the kinetic ligand-binding properties of oxygen, carbon monoxide and nitric oxide to the chimaeric proteins, FHPg (truncated form of FHP comprising the globin domain alone) and VHb-Red (fusion protein between VHb and the C-terminal reductase domain of FHP) and compared them with those of their natural counterparts, FHP and VHb. Moreover, we also analysed polarity and solvent accessibility to the haem pocket of these proteins. The rate constants for the engineered proteins, VHb-Red and FHPg, do not differ significantly from those of their natural counterparts, VHb and FHP respectively. Our results suggest that the globin domain structure controls the reactivity towards oxygen, carbon monoxide and nitric oxide. The presence or absence of a reductase domain does not affect the affinity to these ligands.
Farrés
Judith
J
Institute of Biotechnology, ETH Zürich, CH 8093 Zürich, Switzerland.
Burckhardt-Herold
Susanna
S
Scherrer
Jan
J
Frey
Alexander D
AD
Kallio
Pauli T
PT
D001426:Bacterial Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D001665:Binding Sites; D002248:Carbon Monoxide / Q000737:chemistry / Q000378:metabolism; D020118:Cupriavidus necator / Q000378:metabolism; D006420:Hemeproteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D006454:Hemoglobins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D007700:Kinetics; D008024:Ligands; D009569:Nitric Oxide / Q000737:chemistry / Q000378:metabolism; D010088:Oxidoreductases / Q000737:chemistry* / Q000378:metabolism; D010100:Oxygen / Q000737:chemistry / Q000378:metabolism; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification; D012997:Solvents / Q000378:metabolism; D054793:Truncated Hemoglobins
D001426:Bacterial Proteins; D006420:Hemeproteins; D006454:Hemoglobins; D008024:Ligands; D011993:Recombinant Fusion Proteins; D012997:Solvents; D054793:Truncated Hemoglobins; C086193:flavohemoprotein, Bacteria; C082613:hemoglobin protein, Vitreoscilla; D009569:Nitric Oxide; D002248:Carbon Monoxide; D010088:Oxidoreductases; D010100:Oxygen