TY - JOUR
T1 - Demonstration that histidine-25, but not histidine-132, is the axial heme ligand in rat heme oxygenase-1
AU - Itomaki, Mariko
AU - Ishikawa, Kazunobu
AU - Matera, Kathryn Mansfield
AU - Sato, Michihiko
AU - Ikeda-saito, Masao
AU - Yoshida, Tadashi
N1 - Copyright:
Copyright 2017 Elsevier B.V., All rights reserved.
PY - 1995/2/20
Y1 - 1995/2/20
N2 - A truncated, soluble rat heme oxygenase-1 lacking its C-terminal, membrane-anchoring segment, and its His25 → Ala and His132 → Ala mutants have been prepared by site-directed mutagenesis and expression in Escherichia coli. We found that wild-type enzyme can degrade heme to biliverdin, but its specific activity was about one-fifth that of the native, full-length enzyme, suggesting that the C-terminal segment is important for accepting electrons from NADPH cytochrome P450 reductase. His132 → Ala mutant had an enzyme activity comparable to that of the wild-type enzyme; hence, the highly conserved His132 is not essential for the display of the heme oxygenase activity. In contrast, His25 → Ala mutation completely abolished the enzyme′s catalytic activity. A five-coordinate type ferrous NO EPR spectrum was observed for the heme-heme oxygenase H25A complex. Hence, we conclude that His25 is the proximal axial ligand of the heme iron and is essential for the heme degradation activity of the enzyme.
AB - A truncated, soluble rat heme oxygenase-1 lacking its C-terminal, membrane-anchoring segment, and its His25 → Ala and His132 → Ala mutants have been prepared by site-directed mutagenesis and expression in Escherichia coli. We found that wild-type enzyme can degrade heme to biliverdin, but its specific activity was about one-fifth that of the native, full-length enzyme, suggesting that the C-terminal segment is important for accepting electrons from NADPH cytochrome P450 reductase. His132 → Ala mutant had an enzyme activity comparable to that of the wild-type enzyme; hence, the highly conserved His132 is not essential for the display of the heme oxygenase activity. In contrast, His25 → Ala mutation completely abolished the enzyme′s catalytic activity. A five-coordinate type ferrous NO EPR spectrum was observed for the heme-heme oxygenase H25A complex. Hence, we conclude that His25 is the proximal axial ligand of the heme iron and is essential for the heme degradation activity of the enzyme.
KW - BiJiverdin
KW - EPR
KW - Heme
KW - Heme oxygenase
KW - Hemoprotein
KW - O activation
KW - P450 reductase
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U2 - 10.1006/abbi.1995.1160
DO - 10.1006/abbi.1995.1160
M3 - Article
C2 - 7872792
AN - SCOPUS:0028909671
VL - 317
SP - 253
EP - 258
JO - Archives of Biochemistry and Biophysics
JF - Archives of Biochemistry and Biophysics
SN - 0003-9861
IS - 1
ER -