Determination of iron-carbon monoxide geometry in the subunits of carbonmonoxy hemoglobin M Boston using femtosecond infrared spectroscopy

Abstract

We have undertaken ultrafast infrared (IR) spectroscopic studies in order to elucidate the geometry of bound CO in the alpha and beta subunits of hemoglobin (Hb) M Boston 13CO. Hb M Boston is a mutant human Hb in which the distal histidine in the alpha subunits is replaced by a tyrosine. The IR absorptions of bound 13CO fall at 1925 cm-1 for the alpha subunits and 1907 cm-1 for the beta subunits. Despite a difference of nearly 20 cm-1 in these peaks, the measured anisotropies of the bound 13CO depletions following 30% photolysis are nearly identical, with values of -0.142 +/- 0.002 obtained for the alpha subunits and -0.140 +/- 0.003 obtained for the beta subunits. These translate to values of 20 degrees +/- 1 degree and 21 degrees +/- 1 degree for the values of the average angles between the CO bond and the normal to the heme planes in the alpha and beta subunits, respectively. Our present results and the work of previous investigators [Nagai, M., Yoneyama, Y., & Kitagawa, T. (1991) Biochemistry 30, 6495-6503] suggest that a change in the polar interactions of the bound CO with the heme pocket environment upon substitution of tyrosine for the distal histidine and a less bent structure for the Fe-C-O unit in the alpha subunits are responsible for the difference in the bound CO absorption frequencies in the alpha and beta subunits. A spectrum of the depletion of the bound 13CO peaks following photolysis indicates that both subunits photodissociate CO with the same quantum yield and neither subunit exhibits significant recombination within 1 ns.

Theoretical Study of the Electrostatic and Steric Effects on the Spectroscopic Characteristics of the Metal-Ligand Unit of Heme Proteins. 2. C-O Vibrational Frequencies, 17O Isotropic Chemical Shifts, and Nuclear Quadrupole Coupling Constants

“…This conclusion was consistent with the results of a number of recent experimental studies (Ricard et al, 1986;Kim et al, 1989;Kim and Ibers, 1991;Cameron et al, 1993;Quillin et al, 1993Quillin et al, , 1995Lian et al, 1993;Hu et al, 1994;Ivanov et al, 1994;Ray et al, 1994;Springer et al, 1994;Tetreau et al, 1994;Lim et al, 1995), in which it was shown that the linear perpendicular coordination of CO in HP(CO)s is weakly distorted, if at all, and the wide variation of vco in different HP(CO)s was as-a. FIGURE 1 Schematic representation of the geometry of the complexes with linear perpendicular (a), bent and tilt (b), coordinated CO without and with (c) point charges Qi, and in the presence of the distal histidine (d). signed (Li and Spiro, 1988;Morikis et al, 1989;Augspurger et al, 1991;Oldfield et al, 1991;Park et al, 1991;Brantley et al, 1993;Lian et al, 1993;Cameron et al, 1993;Sakan et al, 1993;Hu et al, 1994;Ivanov et al, 1994;Ray et al, 1994;Springer et al, 1994;Decatur and Boxer, 1995) to the effect of the protein electric field (Geissinger et al, 1995).…”

The Orientation of CO in Heme Proteins Determined by Time-Resolved Mid-IR Spectroscopy: Anisotropy Correction for Finite Photolysis of an Optically Thick Sample

“…Earlier polarized IR measurements on various heme proteins at ambient temperature [21][22][23]25,26,28,29 reported that the orientation of CO in heme protein is bent at different angles ranging from 15 o to 35 o , similar to earlier crystal structure. 5 However, using the same polarized IR spectroscopy, we have recently found that CO in Mb is almost linear, bent less than 7 o relative to the heme plane normal, 15 consistent with more recent higher resolution X-ray crystal structure 16 and spectroscopic data in solution, 30 thereby establishing necessity of newer paradigm for how Mb discriminate against CO.…”

“…The method of photoselection 18,19 combined with polarized IR spectroscopy 15,[20][21][22][23][24][25][26][27][28][29] has been used to determine the orientation of CO relative to the heme plane normal in Mb. Earlier polarized IR measurements on various heme proteins at ambient temperature [21][22][23]25,26,28,29 reported that the orientation of CO in heme protein is bent at different angles ranging from 15 o to 35 o , similar to earlier crystal structure.…”

Theoretical study of the distal-side steric and electrostatic effects on the vibrational characteristics of the FeCO unit of the carbonylheme proteins and their models

“…Third, the electrostatic interaction between the protein polar groups and the heme is under intensive discussion now (Li and Spiro, 1988;Morikis et al, 1989;Augspurger et al, 1991;Oldfield et al, 1991;Park et al, 1991;Brantley et al, 1993;Lian et al, 1993;Cameron et al, 1993;Sakan et al, 1993;Hu et al, 1994;Ivanov et al, 1994; Ray et al, 1994;Springer et al, 1994; Decatur and Boxer, 1995).…”