TY - JOUR
T1 - Binding of oxygen and carbon monoxide to a heme-regulated phosphodiesterase from Escherichia coli
T2 - Kinetics and infrared spectra of the full-length wild-type enzyme, isolated PAS domain, and MET-95 mutants
AU - Taguchi, Sue
AU - Matsui, Toshitaka
AU - Igarashi, Jotaro
AU - Sasakura, Yukie
AU - Araki, Yasuyuki
AU - Ito, Osamu
AU - Sugiyama, Shunpei
AU - Sagami, Ikuko
AU - Shimizu, Toru
PY - 2004/1/30
Y1 - 2004/1/30
N2 - The heme-regulated phosphodiesterase, Ec DOS, is a redox sensor that uses the heme in its PAS domain to regulate catalysis. The rate of O2 association (kon) with full-length Ec DOS is extremely slow at 0. 0019 μM-1 s-1, compared with >9.5 μM -1 s-1 for 6-coordinated globin-type hemoproteins, as determined by the stopped-flow method. This rate is dramatically increased (up to 16-fold) in the isolated heme-bound PAS domain. Dissociation constants (Kd) calculated from the kinetic parameters are 340 and 20 μM for the full-length wild-type enzyme and its isolated PAS domain, respectively. Mutations at Met-95 in the isolated PAS domain, which may be a heme axial ligand in the Fe(II) complex, lead to a further increase in the kon value by more than 30-fold, and consequently, a decrease in the Kd value to less than 1 Μ. The kon value for CO binding to the full-length wild-type enzyme is also very low (0.00081 μM-1 s -1). The kinetics of CO binding to the isolated PAS domain and its mutants are similar to those observed for O2. However, the K d values for CO are considerably lower than those for O2.
AB - The heme-regulated phosphodiesterase, Ec DOS, is a redox sensor that uses the heme in its PAS domain to regulate catalysis. The rate of O2 association (kon) with full-length Ec DOS is extremely slow at 0. 0019 μM-1 s-1, compared with >9.5 μM -1 s-1 for 6-coordinated globin-type hemoproteins, as determined by the stopped-flow method. This rate is dramatically increased (up to 16-fold) in the isolated heme-bound PAS domain. Dissociation constants (Kd) calculated from the kinetic parameters are 340 and 20 μM for the full-length wild-type enzyme and its isolated PAS domain, respectively. Mutations at Met-95 in the isolated PAS domain, which may be a heme axial ligand in the Fe(II) complex, lead to a further increase in the kon value by more than 30-fold, and consequently, a decrease in the Kd value to less than 1 Μ. The kon value for CO binding to the full-length wild-type enzyme is also very low (0.00081 μM-1 s -1). The kinetics of CO binding to the isolated PAS domain and its mutants are similar to those observed for O2. However, the K d values for CO are considerably lower than those for O2.
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U2 - 10.1074/jbc.M301013200
DO - 10.1074/jbc.M301013200
M3 - Article
C2 - 14612459
AN - SCOPUS:0942276394
VL - 279
SP - 3340
EP - 3347
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
SN - 0021-9258
IS - 5
ER -