caa a22 4500 605507694 CHVBK 20210128100633.0 cr unu---uuuuu 210128e20150701xx s 000 0 eng 10.1007/s00775-015-1263-5 doi (NATIONALLICENCE)springer-10.1007/s00775-015-1263-5 Hydrogen bond donation to the heme distal ligand of Staphylococcus aureus IsdG tunes the electronic structure [Elektronische Daten] [Cheryl Lockhart, Matthew Conger, Dylanger Pittman, Matthew Liptak] Staphylococcus aureus IsdG catalyzes the final step of staphylococcal iron acquisition from host hemoglobin, whereby host-derived heme is converted to iron and organic products. The Asn7 distal pocket residue is known to be critical for enzyme activity, but the influence of this residue on the substrate electronic structure was unknown prior to this work. Here, an optical spectroscopic and density functional theory characterization of azide- and cyanide-inhibited wild type and N7A IsdG is presented. Magnetic circular dichroism data demonstrate that Asn7 perturbs the electronic structure of azide-inhibited, but not cyanide-inhibited, IsdG. As the iron-ligating α-atom of azide, but not cyanide, can act as a hydrogen bond acceptor, these data indicate that the terminal amide of Asn7 is a hydrogen bond donor to the α-atom of a distal ligand to heme in IsdG. Circular dichroism characterization of azide- and cyanide-inhibited forms of WT and N7A IsdG strongly suggests that the Asn7···N3 hydrogen bond influences the orientation of a distal azide ligand with respect to the heme substrate. Specifically, density functional theory calculations suggest that Asn7···N3 hydrogen bond donation causes the azide ligand to rotate about an axis perpendicular to the porphyrin plane and weakens the π-donor strength of the azide ligand. This lowers the energies of the Fe 3d xz and 3d yz orbitals, mixes Fe 3d xy and porphyrin a 2u character into the singly-occupied molecular orbital, and results in spin delocalization onto the heme meso carbons. These discoveries have important implications for the mechanism of heme oxygenation catalyzed by IsdG. SBIC, 2015 Circular dichroism nationallicence Density functional theory nationallicence Heme oxygenase nationallicence Magnetic circular dichroism nationallicence Second-sphere interactions nationallicence Abs : Electronic absorption nationallicence CD : Circular dichroism nationallicence DFT : Density functional theory nationallicence Fe(III)TMP(4-CNPy)2 : Bis(4-cyanopyridine)ferrictetramesitylporphyrinate nationallicence Isd : Iron-regulated surface determinant nationallicence IsdG-heme : Heme-bound IsdG nationallicence IsdG-heme-CN : Cyanide-inhibited IsdG nationallicence IsdG-heme-N3 : Azide-inhibited IsdG nationallicence IsdG-heme-OOH : Hydroperoxo-ligated, heme-bound IsdG nationallicence IsdI-heme-OH : Hydroxide-ligated, heme-bound IsdI nationallicence KPi : Potassium phosphate nationallicence MCD : Magnetic circular dichroism nationallicence NaPi : Sodium phosphate nationallicence PMSF : Phenylmethanesulfonyl fluoride nationallicence TEV : Tobacco etch virus nationallicence VTVH : Variable-temperature, variable-field nationallicence WT : Wild-type nationallicence Lockhart Cheryl Department of Chemistry, University of Vermont, 82 University Place, 05405, Burlington, VT, USA aut Conger Matthew Department of Chemistry, University of Vermont, 82 University Place, 05405, Burlington, VT, USA aut Pittman Dylanger Department of Chemistry, University of Vermont, 82 University Place, 05405, Burlington, VT, USA aut Liptak Matthew Department of Chemistry, University of Vermont, 82 University Place, 05405, Burlington, VT, USA aut JBIC Journal of Biological Inorganic Chemistry Springer Berlin Heidelberg 20/5(2015-07-01), 757-770 0949-8257 20:5<757 2015 20 775 https://doi.org/10.1007/s00775-015-1263-5 text/html Onlinezugriff via DOI BK010053 XK010053 XK010000 Metadata rights reserved Springer special CC-BY-NC licence nationallicence 1 research-article jats NATIONALLICENCE NATIONALLICENCE NL-springer NATIONALLICENCE 856 40 https://doi.org/10.1007/s00775-015-1263-5 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Lockhart Cheryl Department of Chemistry, University of Vermont, 82 University Place, 05405, Burlington, VT, USA aut NATIONALLICENCE 700 1- Conger Matthew Department of Chemistry, University of Vermont, 82 University Place, 05405, Burlington, VT, USA aut NATIONALLICENCE 700 1- Pittman Dylanger Department of Chemistry, University of Vermont, 82 University Place, 05405, Burlington, VT, USA aut NATIONALLICENCE 700 1- Liptak Matthew Department of Chemistry, University of Vermont, 82 University Place, 05405, Burlington, VT, USA aut NATIONALLICENCE 773 0- JBIC Journal of Biological Inorganic Chemistry Springer Berlin Heidelberg 20/5(2015-07-01), 757-770 0949-8257 20:5<757 2015 20 775