inorganics Article The Photochemistry of Fe2(S2C3H6)(CO)6(µ-CO) and Its Oxidized Form, Two Simple [FeFe]-Hydrogenase CO-Inhibited Models. A DFT and TDDFT Investigation Federica Arrigoni 1, Giuseppe Zampella 1, Luca De Gioia 1, Claudio Greco 2,* and Luca Bertini 1,* 1 Department of Biotechnology and Biosciences, University of Milano-Bicocca, 20126 Milan, Italy; [email protected] (F.A.); [email protected] (G.Z.); [email protected] (L.D.G.) 2 Department of Earth and Environmental Sciences, University of Milano-Bicocca, 20126 Milan, Italy * Correspondence: [email protected] (C.G.); [email protected] (L.B.) I I 1 I II 2 + Abstract: Fe Fe Fe2(S2C3H6)(CO)6(µ-CO) ( a–CO) and its Fe Fe cationic species ( a –CO) are the simplest model of the CO-inhibited [FeFe] hydrogenase active site, which is known to undergo CO photolysis within a temperature-dependent process whose products and mechanism are still a matter of debate. Using density functional theory (DFT) and time-dependent density functional theory (TDDFT) computations, the ground state and low-lying excited-state potential energy surfaces (PESs) of 1a–CO and 2a+–CO have been explored aimed at elucidating the dynamics of the CO photolysis 1 + 2 + yielding Fe2(S2C3H6)(CO)6 ( a) and [Fe2(S2C3H6)(CO)6] ( a ), two simple models of the catalytic site of the enzyme. Two main results came out from these investigations. First, a–CO and 2a+–CO are both bound with respect to any CO dissociation with the lowest free energy barriers around 10 kcal mol−1, suggesting that at least 2a+–CO may be synthesized. Second, focusing on the cationic Citation: Arrigoni, F.; Zampella, G.; form, we found at least two clear excited-state channels along the PESs of 2a+–CO that are unbound De Gioia, L.; Greco, C.; Bertini, L. with respect to equatorial CO dissociation. The Photochemistry of Fe2(S2C3H6)(CO)6(µ-CO) and Its Keywords: metal-carbonyl complexes; [FeFe]-hydrogenases; density functional theory; time-dependent Oxidized Form, Two Simple [FeFe]-Hydrogenase CO-Inhibited DFT; organometallic photochemistry Models. A DFT and TDDFT Investigation. Inorganics 2021, 9, 16. https://doi.org/10.3390/inorganics 9020016 1. Introduction In recent times, the study of substituted binuclear carbonyl species has gained vast Academic Editor: Duncan Gregory popularity in the context of bioinorganic chemistry due to the fact that the hydrogenase Received: 5 January 2021 enzymes are currently known for their specificity towards dihydrogen oxidation/evolution Accepted: 4 February 2021 invariably include a binuclear carbonyl-containing moiety in their active site [1]. These en- Published: 9 February 2021 zymes, which encompass either only iron ions as metal cofactors ([FeFe]-hydrogenases), or both nickel and iron ([NiFe]-hydrogenases), have inspired the design and synthesis of Publisher’s Note: MDPI stays neutral a plethora of synthetic models to date [2–4], with diiron models being actually prevalent with regard to jurisdictional claims in in literature. Such prevalence depends not only on the interest raised by the knowledge published maps and institutional affil- that [FeFe]-hydrogenases are extremely efficient [5] but also on the fact that diiron hexacar- iations. bonyls of the general formula Fe2(SR)2(CO)6—which closely resemble the diiron portion of FeFe-hydrogenase active site, see Figure1—had been known for seventy years before the publication of the first X-ray structure of the enzyme [6,7]. The availability of a large number of biomimetic catalysts has proved to be the main asset in the quest for a deeper Copyright: © 2021 by the authors. understanding of hydrogenase chemistry [8]. For example, a biomimetic complex de- Licensee MDPI, Basel, Switzerland. scribed Camara, and Rauchfuss [9] has proved highly valuable to confirm the hypothesis This article is an open access article that H2-binding and splitting in [FeFe]-hydrogenases occur on a single Fe center in the distributed under the terms and active site (the so-called “distal” iron ion, Fe in Figure1)[ 10,11]. Interestingly, the same conditions of the Creative Commons d Fe center is thought to be directly involved also in the enzyme inhibition mediated by Attribution (CC BY) license (https:// carbon monoxide [12–14]. CO inhibition is a key topic that has bearings for the perspective creativecommons.org/licenses/by/ 4.0/). [FeFe]-hydrogenases utilization for industrial purposes, as the contact of the enzyme with Inorganics 2021, 9, 16. https://doi.org/10.3390/inorganics9020016 https://www.mdpi.com/journal/inorganics Inorganics 2021, 9, x FOR PEER REVIEW 2 of 19 Inorganics 2021, 9, 16 2 of 17 by carbon monoxide [12–14]. CO inhibition is a key topic that has bearings for the per- spective [FeFe]-hydrogenases utilization for industrial purposes, as the contact of the en- evenzyme traces with even of CO, traces can completelyof CO, can completely impair hydrogenase impair hydrogenase activity [15 ].activity Still, many [15]. Still, aspects many of theaspects chemistry of the chemistry of CO-inhibited of CO-inhibited [FeFe]-hydrogenases [FeFe]-hydrogenases are far from are being far from fully being understood. fully un- Thisderstood. is true This not is only true for not the only rather for the complex rather photochemistry complex photochemistry occurring occurring at the CO-inhibited at the CO- activeinhibited site active (vide infra) site (vide but also infra) with but reference also with to reference the possibility to the of possibility structural rearrangementsof structural re- occurringarrangements at the occurring active site at inthe concomitance active site in with concomitance CO-binding with [14 ,CO-binding16,17]. Notwithstand- [14,16,17]. ingNotwithstanding such open issues, such biomimeticopen issues, modeling biomimetic has modeling had a relatively has had a limited relatively impact limited so farim- forpact the so elucidationfar for the elucidation of the (photo)chemical of the (photo)chemical processes that processes can occur that after can COoccur inhibition, after CO whichinhibition, is mainly which due isto mainly the small due number to the ofsmall biomimetic number models of biomimetic of the CO-inhibited models of enzymethe CO- describedinhibited enzyme to date. described In any case, to date. theavailability In any case, of the the availability latter [18] of has the stimulated latter [18] has positive stim- feedbackulated positive between feedback experiments between and theory,experiment whichs and allowed theory, relating which the allowed hardness relating of ligands the inhardness biomimetic of ligands models in with biomimetic the stability models of key with stereoelectronic the stability featuresof key stereoelectronic in the latter [18 ,19fea-]. Thetures theory-experiment in the latter [18,19]. interplay The theory-experiment proved relevant also interplay in more proved recent studiesrelevant in also which in more den- sityrecent functional studies theoryin which (DFT) density calculations functional were theory used (DFT) to rationalize calculations the structuralwere used outcomes to ration- ofalize CO-inhibition the structural and outcomes subsequent of reductionCO-inhibit ofion the and enzyme subsequent that gives reduction place to over-saturatedof the enzyme formsthat gives of the place active to siteover-saturated [20]. forms of the active site [20]. Figure 1. The active site of [FeFe]-hydrogenases, generally referred to as the H-cluster; its diiron Figure 1. The active site of [FeFe]-hydrogenases, generally referred to as the H-cluster;+ its diiron portion “[2Fe]H” directly involved in the binding of incipient substrates (H2 and H ) and inhibitors + isportion highlighted. “[2Fe]H The” directly carbonyl involved ligand in marked the bind withing of an incipient asterisk substrates represents (H an2 and exogenous H ) and CO inhibitors ligand is highlighted. The carbonyl ligand marked with an asterisk represents an exogenous CO ligand behaving as an inhibitor. A cysteinyl sulfur bridges the diiron subsite with a [Fe4S4] subsite that com- behaving as an inhibitor. A cysteinyl sulfur bridges the diiron subsite with a [Fe4S4] subsite that pletes the H-cluster composition. The two Fe atoms of the diiron subsite are labeled with subscripts completes the H-cluster composition. The two Fe atoms of the diiron subsite are labeled with sub- “d” (distal) or “p” (proximal), depending on their position with respect to the [Fe S ] subsite. scripts “d” (distal) or “p” (proximal), depending on their position with respect to4 the4 [Fe4S4] sub- site. As far as the photochemistry of the CO inhibited form of the enzyme is concerned, it wasAs found far as tothe be photochemistry light-sensitive of at the cryogenic CO inhi temperature.bited form of COthe enzyme photolysis is concerned, is a typical it organometallic light-driven process [21], and this type of temperature-dependent mecha- was found to be light-sensitive at cryogenic temperature. CO photolysis is a typical or- nism has been already observed in Fe (CO) photolysis, as pointed out by Chen et al. [22]. ganometallic light-driven process [21],2 and9 this type of temperature-dependent mecha- In this case, the structure of the Fe2(CO)8 photoproduct depends on the reaction condition: nism has been already observed in Fe2(CO)9 photolysis, as pointed out by Chen et al. [22]. photolysis up to 35 K yields the Fe2(CO)8 bridged form, while at the higher temperature, In this case, the structure of the Fe2(CO)8 photoproduct depends on the reaction condition: the unbridged form is obtained. photolysis up to 35 K yields the Fe2(CO)8 bridged form, while at the higher temperature, This photolytic process is temperature-dependent and has been studied by EPR [23] the unbridged form is obtained. and IR spectroscopy [22,24]. At low temperature (6–8 K), the initial axial EPR signal This photolytic process is temperature-dependent and has been studied by EPR [23] of the CO inhibited form is converted to that of the active form in the absence of CO.