Arylsulfatases require a maturating enzyme to perform a co-or posttranslational modification to form a catalytically essential formylglycine (FGly) residue. In organisms that live aerobically, molecular oxygen is used enzymatically to oxidize cysteine to FGly. Under anaerobic conditions, S-adenosylmethionine (AdoMet) radical chemistry is used. Here we present the structures of an anaerobic sulfatase maturating enzyme (anSME), both with and without peptidyl-substrates, at 1.6-1.8 Å resolution. We find that anSMEs differ from their aerobic counterparts in using backbone-based hydrogen-bonding patterns to interact with their peptidylsubstrates, leading to decreased sequence specificity. These anSME structures from Clostridium perfringens are also the first of an AdoMet radical enzyme that performs dehydrogenase chemistry. Together with accompanying mutagenesis data, a mechanistic proposal is put forth for how AdoMet radical chemistry is coopted to perform a dehydrogenation reaction. In the oxidation of cysteine or serine to FGly by anSME, we identify D277 and an auxiliary [4Fe-4S] cluster as the likely acceptor of the final proton and electron, respectively. D277 and both auxiliary clusters are housed in a cysteinerich C-terminal domain, termed SPASM domain, that contains homology to ∼1,400 other unique AdoMet radical enzymes proposed to use [4Fe-4S] clusters to ligate peptidyl-substrates for subsequent modification. In contrast to this proposal, we find that neither auxiliary cluster in anSME bind substrate, and both are fully ligated by cysteine residues. Instead, our structural data suggest that the placement of these auxiliary clusters creates a conduit for electrons to travel from the buried substrate to the protein surface.iron-sulfur cluster fold | radical SAM dehydrogenase

The bimetallic complexes CpRu(P-P)X [Cp=η 5 -C 5 H 5 ; X=Cl, H; P-P=dppf (1,1 0 -bis(diphenylphosphino)ferrocene), dppr (1,1 0 -bis(diphenylphosphino)ruthenocene), dppo (1,1 0 -bis(diphenylphosphino)-osmocene), dippf (1,1 0 -bis(diisopropylphosphino)ferrocene), dcpf (1,1 0 -bis(dicyclohexylphosphino)ferrocene)], Cp*Ru(P-P)X [Cp*=η 5 -C 5 Me 5 ; X=Cl, H; P-P=dppf, dippf, dppomf (1,1 0 -bis(diphenylphosphino) octamethylferrocene), dppc (1,1 0 -bis(diphenylphosphino)cobaltocene)], [Cp*Ru(P-P)X] + (X=H, CCPh; P-P=dppc + ), and [Cp*Ru(P-P)L] 2+ (L= CH 3 CN, t-BuCN; P-P= dppc + ) have been synthesized. Most of the chloride and hydride complexes have been studied by cyclic voltammetry. The X-ray structures of [Cp*Ru(dppc)CH 3 CN][PF 6 ] 2 and [Cp*Ru(dppc)CCPh][PF 6 ] have been determined. Protonation of [Cp*Ru(dppc)CCPh] + gives the vinylidene complex [Cp*Ru(dppc)CCHPh] 2+ . The Co(III/II) potential of the dppc + ligand undergoes a cathodic shift upon coordination in [Cp*Ru-(dppc)H] + and an anodic shift upon coordination in [Cp*Ru(dppc)CH 3 CN] 2+ . The 1 H NMR spectrum of Cp*Ru(dppc)H is consistent with its formulation as a Co(II)/Ru(II) complex. As gauged by their reactivity toward iminium cations, the hydride complexes are poor hydride donors; proton and electron transfer are dominant. CpRu(dippf)H and CpRu(dcpf)H deprotonate iminium cations with acidic R-hydrogens. Cp*Ru(dppc)H is oxidized by the N-(benzylidene)pyrrolidinium cation, giving [Cp*Ru-(dppc)H] + and the vicinal diamine 1,2-bis(N-pyrrolidino)-1,2-diphenylethane. Most of the hydride complexes give trans-dihydride cations upon protonation; an exception is [Cp*Ru(dppc)H] + , which forms a dihydrogen complex [Cp*Ru(dppc)(H 2 )] 2+ with surprising kinetic stability. This dihydrogen complex is more acidic and less thermodynamically stable than its dihydride isomer. The H 2 ligand in [Cp*Ru(dppc)-(H 2 )] 2+ is readily replaced by nitriles; the reaction with t-BuCN occurs by a dissociative mechanism.

No abstract

No abstract