Chemistry and Molecular Dynamics Simulations of Heme b-HemQ and Coproheme-HemQ.
Recently, a novel pathway for heme b biosynthesis in Gram-positive bacteria has been proposed. The final poorly understood step is catalyzed by an enzyme called HemQ and includes two decarboxylation reactions leading from coproheme to heme b. Coproheme has been suggested to act as both substrate and redox active cofactor in this reaction. In the study presented here, we focus on HemQs from Listeria monocytogenes (LmHemQ) and Staphylococcus aureus (SaHemQ) recombinantly produced as apoproteins in Escherichia coli. We demonstrate the rapid and two-phase uptake of coproheme by both apo forms and the significant differences in thermal stability of the apo forms, coproheme-HemQ and heme b-HemQ. Reduction of ferric high-spin coproheme-HemQ to the ferrous form is shown to be enthalpically favored but entropically disfavored with standard reduction potentials of -205 ± 3 mV for LmHemQ and -207 ± 3 mV for SaHemQ versus the standard hydrogen electrode at pH 7.0. Redox thermodynamics suggests the presence of a pronounced H-bonding network and restricted solvent mobility in the heme cavity. Binding of cyanide to the sixth coproheme position is monophasic but relatively slow (∼1 × 10(4) M(-1) s(-1)). On the basis of the available structures of apo-HemQ and modeling of both loaded forms, molecular dynamics simulation allowed analysis of the interaction of coproheme and heme b with the protein as well as the role of the flexibility at the proximal heme cavity and the substrate access channel for coproheme binding and heme b release. Obtained data are discussed with respect to the proposed function of HemQ in monoderm bacteria.
Department for Structural and Computational Biology, Max F. Perutz Laboratories, University of Vienna , A-1030 Vienna, Austria.
Stefan
S
Hofbauer
Department of Material Sciences and Process Engineering, Institute of Molecular Modeling and Simulation, BOKU-University of Natural Resources and Life Sciences , A-1190 Vienna, Austria.
Marco
M
Dalla Sega
Department of Food Science and Technology, Food Biotechnology Laboratory, BOKU-University of Natural Resources and Life Sciences , A-1190 Vienna, Austria.
Stefan
S
Scheiblbrandner
Department of Material Sciences and Process Engineering, Institute of Molecular Modeling and Simulation, BOKU-University of Natural Resources and Life Sciences , A-1190 Vienna, Austria.
Zuzana
Z
Jandova
Department of Chemistry, Division of Biochemistry, VIBT-Vienna Institute of BioTechnology, BOKU-University of Natural Resources and Life Sciences , A-1190 Vienna, Austria.
Irene
I
Schaffner
Department for Structural and Computational Biology, Max F. Perutz Laboratories, University of Vienna , A-1030 Vienna, Austria.
Georg
G
Mlynek
Department for Structural and Computational Biology, Max F. Perutz Laboratories, University of Vienna , A-1030 Vienna, Austria.|Department of Biochemistry, Faculty of Chemistry and Chemical Technology, University of Ljubljana , 1000 Ljubljana, Slovenia.
Kristina
K
Djinović-Carugo
Department of Chemistry and Geology, University of Modena and Reggio Emilia , 41125 Modena, Italy.
Gianantonio
G
Battistuzzi
Department of Chemistry, Division of Biochemistry, VIBT-Vienna Institute of BioTechnology, BOKU-University of Natural Resources and Life Sciences , A-1190 Vienna, Austria.
Paul G
PG
Furtmüller
Department of Material Sciences and Process Engineering, Institute of Molecular Modeling and Simulation, BOKU-University of Natural Resources and Life Sciences , A-1190 Vienna, Austria.
Chris
C
Oostenbrink
Department of Chemistry, Division of Biochemistry, VIBT-Vienna Institute of BioTechnology, BOKU-University of Natural Resources and Life Sciences , A-1190 Vienna, Austria.
Christian
C
Obinger
D002942:Circular Dichroism; D004578:Electron Spin Resonance Spectroscopy; D006418:Heme / Q000737:chemistry*; D007700:Kinetics; D008024:Ligands; D056004:Molecular Dynamics Simulation; D010802:Phylogeny; D013056:Spectrophotometry, Ultraviolet
D008024:Ligands; D006418:Heme