Proton NMR spectroscopy of sulfmyoglobin

Abstract

The 1H NMR spectra of ferrous sulfmyoglobin, metsulfmyoglobin, and ferric cyanosulfmyoglobin were obtained at 300 MHz. Hyperfine-shifted resonances are observed in the case of metsulfmyoglobin and ferric cyanosulfmyoglobin that have line widths and cover a chemical shift range that are comparable to the corresponding forms of normal myoglobin. Two methyl resonances are observed in the spectrum of ferric cyanosulfmyoglobin at 44.19 and 25.48 ppm (25 degrees C, pH 8.3) that have been assigned to heme methyls at the 8- and 5-positions on the basis of pH titration effects homologous to the corresponding methyl resonances in ferric cyanomyoglobin. Examination of aromatic region resonances and the pH titration profiles of histidine resonances lead to the conclusion that the overall conformation of sulfmyoglobin was highly homologous to that of normal myoglobin and afforded assignments of histidine residues of the former. The most likely position for the addition of a sulfur atom to the heme of sulfmyoglobin is pyrrole ring A, with ring B a possible, but less likely, alternative.

Charge Transfer and π to π* Transitions in the Visible Spectra of Sulfheme Met Isomeric Structures

“…NMR experiments indicate that the 4-vinyl group of the heme is not necessary for the initial formation of the S A sulfheme isomer, though clearly this group is required for formation of the S C sulfheme isomer. [20][21][22][23][24]28,61,62 Thus, the electronic properties of the sulfheme adduct, specifically the changes in the pyrrolic β ring, have a considerable effect on the energy of the heme HOMO orbitals modulating the separation energy between the a 1u and a 2u orbitals. 60 S A , S B , and S C Sulfheme Electronic Excitations.…”

“…Subsequent studies demonstrated that oxygen binding to sulfMb and to sulfHb was reduced, in relation to the normal heme proteins, by ∼2500 and ∼135 times, respectively. − The presence of an analogous sulfur chlorin ring in sulfheme has been supported by the three-dimensional structure of cyanomet-sulfmyoglobin, resonance Raman, NMR, − hemoglobin I mutants from Lucina pectinata , − catalase, , and lactoperoxidase (LPO) . There are heme proteins that do not form sulfheme such as hemoglobin I (HbI) from L. pectinata , which has Gln64 in the E7 position.…”

Hydrogen sulfide activation in hemeproteins: The sulfheme scenario

“…Michel et al showed the formation of an analogous compound upon interaction of Mb-O 2 with H 2 S and termed the complex sulfmyoglobin (sulfMb) [25–27]. These derivatives have a sulfur atom incorporated in the heme pyrrole B with a characteristic optical band around 620 nm [26, 28–31]. The displacement of the band depends on heme-Fe oxidation and ligation states, as well as the type of the sulfheme isomer [26, 32–34].…”

“…Using labeled sulfur ( 35 S) experiments it was first determined that only one atom of sulfur was incorporated into the heme group of sulfMb [26–28, 44, 46]. Later, proton NMR studies of sulfMb identified the sulfur atom on pyrrole B by analyzing the chemical shifts of the heme 1,3,5,8-methyls and the 2,4-vinyl substitutions [31, 34, 47]. Based on NMR and resonance Raman data it was then proved that the sulfur atom is incorporated across the double β-β bond of pyrrole B either as episulfide or thiochlorin, with a heme-chlorin type structure [26, 28, 30, 31, 34, 44, 47, 48].…”

“…Similar to the native proteins, sulfMb and sulfHb bind numerous ligands at distal site of the heme iron [25, 26, 29, 31–33, 35]. Spontaneous autoxidation as well as oxidation state interconversion with different reducing and oxidation reagents are also maintained in these sulfheme derivatives without disturbing the sulfur ring.…”