Carbon monoxide binding properties of domain-swapped dimeric myoglobin

Abstract

Myoglobin (Mb) is a monomeric oxygen storage hemoprotein, and has been shown to form a domain-swapped dimer. In this study, monomeric and dimeric carbon monoxide (CO)-bound Mb (MbCO) exhibited similar absorption spectra. The CO stretching frequencies of MbCO were observed at 1,932 and 1,944 cm(-1) for both monomeric and dimeric MbCO. The resonance Raman (RR) bands for the stretching between the heme iron and axial ligands were observed at the same frequencies for the monomer and dimer of deoxygenated Mb (deoxyMb) and MbCO, respectively (ν Fe-His, 220 cm(-1); ν Fe-C, 507 cm(-1)), showing that the Fe-His bond strength of deoxyMb and the Fe-CO bond strength of MbCO did not change by the dimerization. Time-resolved RR measurements showed that the dynamics of the structural changes at the heme active site after CO photo-dissociation of MbCO was similar between monomeric and dimeric Mb [monomer, (5.2 ± 1.8) × 10(6) s(-1); dimer, (6.2 ± 1.1) × 10(6) s(-1) at room temperature]. These results show that the heme coordination structure, the protein environment around the bound CO, and the protein relaxation character are similar between monomeric and dimeric MbCO. Although the active site structure was similar between the monomer and dimer, the CO binding rate constant of dimeric Mb [(1.01 ± 0.03) × 10(6) M(-1) s(-1) at 20 °C] was about twice larger than that of the monomer [(0.52 ± 0.02) × 10(6) M(-1) s(-1) at 20 °C], presumably due to the expansion of the channel between the Xe3 cavity and the solvent by the dimerization.

Insights into the Active Site of Coproheme Decarboxylase from Listeria monocytogenes

“… 42 , 43 The position of the distal His has been identified to be responsible for the different forms, which correspond to a closed (A 1 ) and an open form (A 0 ). 43 Unlike SWMb, for HHMb only one species has been identified by RR corresponding to form A 1 , 44 while two ν(CO) stretching modes (at 1944 and 1932 cm –1 ) have been observed in the IR spectrum, 45 suggesting the presence also of form A 3 . By analogy, we name the two conformers observed in the coproheme-Mb complex A 1 (510/1941 cm –1 ) and A 0 (503/1956 cm –1 ), however, this latter form is not completely open but shows only a decreased polar interaction with the distal residues.…”

“…Panel B: experimental conditions: Mb and coproheme: λ exc 406.7 nm, laser power at the sample 5 mW, average of 4 spectra with 40 min integration time and 10 spectra with 100 min integration time in the low and high frequency regions, respectively (Mb), average of 6 spectra with 60 min integration time and 12 spectra with 120 min integration time in the low and high frequency regions, respectively (coproheme); WT and its mutants, λ exc 413.1 nm, laser power at the sample 1−3 mW; average of 28 spectra with 280 min integration time and 22 spectra with 220 min integration time in the low and high frequency regions, respectively (WT), average of 6 spectra with 60 min integration time and 18 spectra with 180 min integration time in the low and high frequency regions, respectively (M149A), average of 6 spectra with 60 min integration time and 15 spectra with 150 min integration time in the low and high frequency regions, respectively (M149A/Q187A), and average of 9 spectra with 90 min integration time and 15 spectra with 150 min integration time in the low and high frequency regions, respectively (Q187A). spectrum, 45 suggesting the presence also of form A 3 . By analogy, we name the two conformers observed in the coproheme-Mb complex A 1 (510/1941 cm −1 ) and A 0 (503/ 1956 cm −1 ), however, this latter form is not completely open but shows only a decreased polar interaction with the distal residues.…”

Oligomerization of cytochrome c, myoglobin, and related heme proteins by 3D domain swapping

“…Although the dimer active site was formed from two Mb protomers with the proximal and distal sites of the heme originating from different protomers, the dimer exhibited high O2 binding capacity, similar to monomeric Mb. The protein environment around the bound CO and the protein relaxation character were also similar between the CO-bound Mb monomer and dimer [71]. However, the CO binding rate constant of the Mb dimer ((1.01 ± 0.03) × 10 6 M -1 s -1 at 20°C) was about twice as large as that of the monomer ((0.52 ± 0.02) × 10 6 M -1 s -1 at 20°C), presumably due to the expansion of the channel between the Xe3 cavity [72] and the solvent by the dimerization.…”

Probing Structure and Reaction Dynamics of Proteins Using Time-Resolved Resonance Raman Spectroscopy

“…However, a small fraction of dimers exist in conventional preparations, which can separate from the monomeric fraction with chromatography . Such domain-swapped Mb dimers have been isolated and studied with respect to their CO binding behavior, using ns TR RR experiments . Both the frequencies of the characteristic porphyrin modes and their temporal evolution do not display any significant differences compared to monomeric Mb, which further supports the view of the relatively weak effect of the protein on the structure and dynamics of the heme.…”