The First Example of a Nitrile Hydratase Model Complex that Reversibly Binds Nitriles

Abstract

Nitrile hydratase (NHase) is an iron-containing metalloenzyme that converts nitriles to amides. The mechanism by which this biochemical reaction occurs is unknown. One mechanism that has been proposed involves nucleophilic attack of an Fe-bound nitrile by water (or hydroxide). Reported herein is a five-coordinate model compound ([Fe III (S 2 Me2 N 3 (Et,Pr))] + ) containing Fe(III) in an environment resembling that of NHase, which reversibly binds a variety of nitriles, alcohols, amines, and thiocyanate. XAS shows that five-coordinate [Fe III (S 2 Me2 N 3 (Et,Pr))] + reacts with both methanol and acetonitrile to afford a six-coordinate solvent-bound complex.Competitive binding studies demonstrate that MeCN preferentially binds over ROH, suggesting that nitriles would be capable of displacing the H 2 O coordinated to the iron site of NHase. Thermodynamic parameters were determined for acetonitrile (ΔH = −6.2(±0.2) kcal/mol, ΔS = −29.4(±0.8) eu), benzonitrile (−4.2(±0.6) kcal/mol, ΔS = −18(±3) eu), and pyridine (ΔH = −8(±1) kcal/mol, ΔS = −41(±6) eu) binding to [Fe III (S 2 Me2 N 3 (Et,Pr))] + using variable-temperature electronic absorption spectroscopy. Ligand exchange kinetics were examined for acetonitrile, isopropylnitrile, benzonitrile, and 4-tert-butylpyridine using 13 C NMR line-broadening analysis, at a variety of temperatures. Activation parameters for ligand exchange were determined to be ΔH ‡ = 7.1(±0.8) kcal/mol, ΔS ‡ = −10(±1) eu (acetonitrile), ΔH ‡ = 5.4(±0.6) kcal/mol, ΔS ‡ = −17(±2) eu (iso-propionitrile), ΔH ‡ = 4.9(±0.8) kcal/mol, ΔS ‡ = −20(±3) eu (benzonitrile), and ΔH ‡ = 4.7(±1.4) kcal/mol ΔS ‡ = −18(±2) eu (4-tert-butylpyridine). The thermodynamic parameters for pyridine binding to a related complex, [Fe III (S 2 Me2 N 3 (Pr,Pr))] + (ΔH = −5.9(±0.8) kcal/mol, ΔS = −24(±3) eu), are also reported, as well as kinetic parameters for 4-tert-butylpyridine exchange (ΔH ‡ = 3.1(±0.8) kcal/mol, ΔS ‡ )−25(±3) eu). These data show for the first time that, when it is contained in a ligand environment similar to that of NHase, Fe(III) is capable of forming a stable complex with nitriles. Also, the rates of ligand exchange demonstrate that low-spin Fe(III) in this ligand environment is more labile than expected. Furthermore, comparison of (Tables S-6-S-9), van't Hoff plots for ligand binding to 1 and 2, variable-temperature electronic absorption spectra (Figures S-3-S-7), and EPR spectra for "substrate"-bound 2 , and crystallographic data for 2-NCS (Tables S-10-S-14) (PDF). This material is available free of charge via the Internet at http://pubs.acs.org. HHS Public AccessAuthor manuscript (S 2 Me2 N 3 (Pr,Pr))] + demonstrates how minor distortions induced by ligand constraints can dramatically alter the reactivity of a metal complex.Performing reactions under environmentally friendly conditions has recently become a desirable goal in the search for new catalysts. 1 Enzymes are often viewed as ideal in this respect because of their ability to perform chemical transformations und...

Sulfur K-Edge XAS and DFT Calculations on Nitrile Hydratase: Geometric and Electronic Structure of the Non-heme Iron Active Site

“…[14][15][16][17][18] A significant number of structurally relevant model complexes have been reported, some of which bind nitriles and some of which reversibly bind NO and show photolytic behavior. [19][20][21][22] Furthermore, the study of a series of model complexes having oxidized ligands has shown that this oxidation can significantly alter the pK a of a water bound to the low-spin Fe III . 23 Ligand K-edge X-ray absorption spectroscopy (XAS) is a direct probe of a ligand's chemical nature and its bonding to a metal.…”

Mononuclear, Dinuclear, and Pentanuclear [{N,S(thiolate)}Iron(II)] Complexes: Nuclearity Control, Incorporation of Hydroxide Bridging Ligands, and Magnetic Behavior

“…Zinc, [1][2][3][4][5][6][7] nickel, [8][9][10][11][12] and iron [13][14][15][16][17][18][19][20] thiolate complexes have been made as models for mononuclear and polynuclear metalloenzyme active sites with RSÀM coordination. The alkylthiolate functionality is important because of its relevance to cysteinate ligation, but it is also quite challenging to control from a synthetic perspective.…”

How does cyanide inhibit superoxide reductase? Insight from synthetic Fe III N 4 S model complexes

“…2, the acetate is coordinated in a monodentate, as opposed to bidentate, fashion trans to an imine nitrogen [N(1), Table 1]. As is the case with structures 3 and 5, the FeOS bond in 6 (Table 1) falls on the short end of reported low-spin Fe(III) thiolate structures (21,22,30). Again, this most likely reflects the cationic molecular charge and the weaker trans influence of the amine nitrogen [N(2)] relative to anionic ligands such as N 3 Ϫ or SCN Ϫ .…”

“…4b) are roughly half [as opposed to significantly less (23,30)] as intense as the oscillations due to first coordination sphere atoms (rЈ between 1 and 2 Å). Because scattering by C is not significantly different from that by N, we initially analyzed the first sphere (rЈ ϭ 0.8-2.0 Å) Fourier filtered (FF 1 ) EXAFS by using a single S scatterer and five N scatterers; the latter were distributed among one or more shells, each with an integral n (no.…”

“…Then the Fourier transform range was expanded (rЈ ϭ 0.8-3.0 Å) to include the second-sphere EXAFS (FF 12 ), and additional shells were added (with a common refined second sphere 2 ) to fit the data. The best fit to the FF 12 ( 2 ϭ 1.2) required three additional shells, the cyano N (with multiple scattering effects included) and two C shells at 2.9 and 3.4 Å [the latter is thought to model ligand-based multiple scattering pathways (30)]. Omission of any one of these three shells gave a much worse fit ( 2 Ͼ 3).…”