Vibrational spectroscopic studies of an [Fe8S9]-cluster containing reductase and its ATP-dependent redox partner

ULISES A. ZITARE; SAGIE KATZ; KONSTANTIN LAUN; CHRISTIAN LORENT; JAE-HUN JEOUNG; MARIA ANDA MROGINSKY; HOLGER DOBBEK; INGO ZEBGER

Grenoble

Congreso; 16th European Biological Inorganic Chemistry Conference (EuroBIC-16); 2022

SBIC

Demanding reductive reactions in biology typically depend on ATPases harboring FeS-cluster/s. These enzymes usually constitute a two-component system where ATP hydrolysis in the donor protein is coupled to a redox reaction, providing the energy an electron needs to reduce the metal site on the acceptor protein that catalyzes the reduction. Herein, we present an IR and resonant Raman (RR) spectroscopic characterization of such a two-component system bearing a [Fe8S9]-cluster [{Fe4S4(SCys)3}2(μ2-S)] as active site. This recently discovered double cubane cluster (DCC) is the first example in nature so far.1 The DCC-containing protein (DCCP) forms a complex with its specific reductase (DCCP-R) that is capable of performing efficient two-electron chemistry at very low redox potentials. Thereby, acetylene (C2H2) and other small molecules as azide (N3 −) or hydrazine (N2H4) are reduced.Frozen solution samples and crystals of DCCP, DCCP-R and the entire complex were characterized by resonant Raman spectroscopy after different redox treatments. Supporting calculations allowed the identification of a vibrational mode involving a single sulfur bridging both cubanes, which is suggested to be the key atom for the reduction of small molecule.Scaffold rearrangements involved in the catalytic mechanism were studied by transmission IR in a thin layer cell employing caged nucleotides. Difference spectra upon ATP release revealed a well-defined band pattern in the amide I region that was tentatively assigned to specific secondary structure elements and amino acid residues. Notably, ATP binding (and not hydrolysis) triggers the conformational change and time resolved measurements suggest that this might be the gating step for the entire catalytic process. This ongoing vibrational spectroscopic study of the DCCP/DCCP-R complex sheds light on its catalytic mechanism and broadens our understanding of the specifics of a new Fe/S-cluster.