Copper(I) Nitrosyls from Reaction of Copper(II) Thiolates with

Copper(I)Nitrosyls from Reaction of Copper(II)Thiolates with S ?Nitrosothiols:Mechanism of NO Release from RSNOs at Cu

Shiyu Zhang,?Nihan C e lebi-O lc u m ,*,§,∥Marie M.Melzer,?,?K.N.Houk,§and Timothy H.Warren ?,*?Department of Chemistry,Georgetown University,Box 571227-1227,Washington,DC 20057,United States

§Department of Chemistry and Biochemistry,University of California,Los Angeles,California 90095-1569

∥Department of Chemical Engineering,Yeditepe University,Istanbul 34755,Turkey

*Supporting Information

S -nitrosothiols,RSNOs,serve both as ready reservoirs of NO activity and active agents in the post-translational modi ?cation of proteins through S -transnitrosation of cysteine SH residues.1Low-molecular weight RSNOs such as S -nitrosoglutathione (GSNO)are present at submicromolar concentrations in the plasma 2and exhibit a number of bene ?cial physiological e ?ects 1such as protection against myocardium 3and lung/airway 3injuries.Due to the modest RS-NO bond strength (20?32kcal/mol),4RSNOs serve as air-stable reservoirs of NO.O ?ering the prospect of controlled,triggered NO release in biological environments,copper ions are e ?ective in catalyzing RSNO decomposition to NO gas and disul ?des RSSR (Scheme

1).5CuZnSOD which represents the most abundant source of copper in red blood cells releases NO from GSNO.6NO release may be inhibited with neocuproine,a Cu +-speci ?c chelator.6Taking advantage of endogenous RSNOs in the plasma,Cu 2+ions embedded into medical polymers can serve as long-lived NO generating devices.7Thus,copper complexes starting in both the Cu(I)and Cu(II)oxidation states can be e ?ective for release of NO from RSNOs.We recently reported the formation of NO in the reaction of

an electron-rich β-diketiminato copper(I)complex [Me 2NN]Cu with Ph 3CSNO to give the copper(II)thiolate

[Me 2NN]Cu ?SCPh 3(Scheme 1).8Release of NO from E ?NO at a Cu(I)center is the microscopic reverse of reductive nitrosylation,which occurs at Cu(II)centers [Cu II ]?E with concomitant

formation of the organonitroso compound E ?

NO.9Related reductive nitrosylation reactions involving the

formation of O ?NO bonds has been observed in laccase 10and

other enzymes.11The formation of N ?NO bonds with NO is a

common pathway in reaction of copper(II)amine complexes 12

and has been used as the basis for NO detection.13

Copper(II)thiolates are ubiquitous in biology,especially in “

blue copper ”type 1Cu sites possessing strong absorbances near λ=600nm (ε≈3000?4000M ?1cm ?1)14that mediate electron transfer in a number of enzymes.Ceruloplasmin,the most

abundant source of copper in the plasma (～1.5?3.1μM),15contains three type 1Cu sites,two with Met donors and one without.Coupled with the relatively high concen-

tration of low molecular weight S -nitrosothiols (0.2?0.3μM)in the plasma,2the possibility of interaction between copper(II)thiolates related to type 1Cu with RSNOs is of interest,

especially since ceruloplasmin has been shown to generate RSNOs from NO and thiols such as glutathione.16

In this study,we employ copper(II)thiolates supported by tris(pyrazolyl)borates,which have been used as structural 17and

spectroscopic 17,18models for type 1Cu sites.These biological copper centers feature two histidine N-donors,an anionic cysteine S-donor in addition to a weak donor such as methionine that results in only a slightly distorted trigonal environment at copper.X-ray structures of TpCu ?SR species generally reveal two shorter Cu ?N (1:1.930(9),2.037(9);17a 2:1.97(5),2.03(4))17b and one modestly longer Cu ?N (1:2.119(8);2:2.05(4))distances with a relatively short Cu ?S distance (1:2.176(4);2:2.12(2)).

Received:June 28,2013

Scheme 1.Copper-Mediated Reactions with S

-Nitrosothiols

To examine the reaction between copper(II)thiolates and RSNOs,we employed the previously reported iPr2TpCu ?SR (R =C 6F 5(1)and CPh 3(2))which possess reasonable thermal stability.Reaction of 1with C 6F 5SNO (prepared in situ from [NO]BF 4and TlSC 6F 5)in CH 2Cl 2(at 0°C over 12min)led to partial consumption of 1(12%)as judged by the loss of the strong band of 1at λmax =666nm (ε≈5900M ?1cm ?1)with formation of a new band at λmax =495nm.GC/MS and 19F NMR analysis indicates formation of the disul ?de C 6F 5S ?

SC 6F 5in 18%yield (Scheme 2).Addition of the much larger S -nitrosothiol Ph 3CSNO to the bulky copper(II)thiolate iPr2Tp-Cu ?SCPh 3(λmax =625nm;ε≈6600M ?1cm ?1)gave no reaction under similar conditions.The new optical band at λmax =495nm in the reaction of 1with C 6F 5SNO is the copper(I)nitrosyl iPr2TpCu(NO)(3)which may be generated independently by reaction of iPr2TpCu with excess NO (λmax =495nm (ε≈960M ?1cm ?1));νNO =1704cm ?1)at ?40°C.While it is clear that iPr2TpCu(NO)(3)forms in the reaction of iPr2TpCu ?SC 6F 5and C 6F 5SNO,3is di ?cult to quantify because of its relative instability.Related tris(pyrazolyl)borate copper(I)nitrosyls TpCu(NO)feature labile binding of NO which may also disproportionate over time to TpCu(NO 2)and N 2O.19Nonetheless,addition of the disul ?de C 6F 5S ?SC 6F 5to iPr2TpCu(NO)(3)(in situ generated at ?40°C in CH 2Cl 2)at 0°C results in the formation of iPr2TpCu ?SC 6F 5(1)(68%yield;UV ?vis)and C 6F

5SNO,

indicating an equilibrium (Scheme 2).Monitoring over the temperature range ?70°C to ?40°C gives the thermodynamic parameters ΔH r =?2.3(2)kcal/mol and ΔS r =?14.5(8)for this reversible transformation (Figure S5).To better understand the fate of the S -residues in the reaction of iPr2TpCu ?SR with S -nitrosothiols,we added the distinct S -nitrosothiol t BuSNO to 1and 2at 25°C in CH 2Cl 2(Scheme 3).In each case,the predominant S-containing product is the unsymmetrical disul ?de RS ?S t Bu.Following the reaction of 1equiv t BuSNO to iPr2TpCu ?SC 6F 5(1)by UV ?vis spectroscopy results in the decay of 1and formation of iPr2TpCu(NO)(3).19F NMR analysis following reaction in benzene-d 6reveals that the unsymmetrical disul ?de C 6F 5S ?S t Bu is the sole ?uorine-containing product;the disul ?de t BuS ?S t Bu accounts for the remainder of the t BuSNO employed.The reaction between iPr2TpCu ?SCPh 3(2)and t BuSNO proceeds similarly with loss of 2,formation of 3,and identi ?cation of Ph 3CS ?S t Bu as the major new S-containing product with t BuS ?S t Bu as the remainder.A preliminary kinetic study via initial rates identi ?es that the reaction is clearly ?rst order in t BuSNO when the initial iPr2TpCu ?SCPh 3concentration is held constant (Scheme S4).The absence of the symmetric disul ?des C 6F 5S ?SC 6F 5and Ph 3CS ?SCPh 3(<5%)in these reactions suggested that S -transnitrosation does not compete with Cu ?NO and RS ?SR bond formation.This is in contrast to the closely related zinc thiolates iPr2TpZn-SR ′which react with RSNO to cleanly undergo S -transnitrosation to give equilibrium mixtures of iPr2TpZn-SR and R ′SNO (Scheme 4).20The later reaction appears to be a zinc-mediated transnitrosation of thiolate anions R ′S ?with S -nitrosothiols RSNO

which have been shown by experiment 21a and theory 21b to proceed via nitroxyl disul ?de intermediates [R ′S(NO)SR]?(Scheme 4).

We used

dispersion-corrected density functional theory (DFT-D3)22and Gaussian 0923to investigate the reactivity of Cu(II)thiolates with S -nitrosothiols.For computational e ?ciency,we considered iPr TpCu ?SC

6F 5,a steric model of 1that involves only the isopropyl substituent on each pyrazolyl ring that ?anks the coordination pocket.Optimization results in two short and one long Cu ?N distance (1.99?and 2.24?,respectively)and a Cu ?S distance of 2.22?at the B3LYP/6-311G(d)level,in good agreement with the X-ray structure of 1.Dispersion corrected energies suggest a slightly endothermic reaction between iPr TpCu ?SC

6F 5and C 6F 5SNO (ΔH

r =1.2

kcal/mol)in accordance with the experimentally observed reversibility

of the reaction (Scheme 2).The reaction of iPr TpCu ?SC

6F 5with t BuSNO is predicted to be slightly

exothermic (ΔH

r =?2.1kcal/mol).These are close to values measured by experiment (?2.3(2)kcal/mol and ?1.7(1)kcal/mol,respectively).

We ?rst considered

the possibility of a nitroxyl disul ?de anion

[RS(NO)SR ′]?bound to a copper(II)center in iPr2TpCu(κ2-SR(NO)SR ′)along the reaction pathway.This

was especially attractive,since theoretical studies on nitroxyl

disul ?des have suggested that these are rather unstable toward oxidation to NO and the corresponding disul ?de (Scheme 4).For instance,the aqueous oxidation potential of [MeS(NO)-SMe]?anion was estimated as +0.31vs NHE 21b and the oxidation potential of iPr2TpCu(NCMe)is +0.64V in MeCN.24

Quantum mechanical investigations on a small model system

employing MeSNO with TpCu ?SMe possessing the unsub-stituted tris(pyrazolyl)borate ligand were performed at the B3LYP-D3/6-311+G(d)level of theory (Scheme 5).Formation of the symmetric nitroxyl disul ?de intermediate TpCu(κ2-MeS(NO)SMe)(4)is predicted to be exothermic with respect to the starting materials (ΔH

r =?14.2kcal/mol and ΔG r =0.7kcal/mol).The optimized structure of 4shown in Scheme 5,however,suggests a sterically unfavorable orientation of the

alkyl groups of the nitroxyl disul ?de toward the Tp ligand,which is partly compensated by favorable dispersion interactions.Indeed,inclusion of the steric bulk around the reaction center using the steric model iPr TpCu revealed that

Scheme 2.Reactivity of iPr2TpCu ?SR with

RSNO Scheme 3.Reactivity of iPr2TpCu ?SR with t BuSNO at 25°

C Scheme 4.Reactivity of Thiolates with S

-nitrosothiols

formation of the corresponding nitroxyl disul ?de 6from iPr TpCu ?SC

6F 5and t BuSNO is sterically more challenging

(ΔH r =?1.1kcal/mol and ΔG r (298K)=12.5kcal/mol;Scheme 6).

We also considered direct attack of an RSNO at the metal center that leads to a Cu ?N interaction in TpCu(SMe)(κ1-N(O)SMe)(Scheme 5).While this κ1-N binding mode for MeSNO at a naked Cu +ion previously has been theoretically considered,25isolable complexes bearing S -nitrosothiols coor-dinated to transition metal ions are exceedingly rare and are limited to a few kinetically inert,low-spin Ir(III)species.26Although κ1-N coordination of MeSNO to TpCu ?SMe to give 5(ΔH r =?4.9kcal/mol,ΔG r =7.4kcal/mol)is predicted to be less favorable than 4,we ?nd that κ1-N coordination of the much bulkier t BuSNO to iPr TpCu ?SC 6F 5in 7is substantially more stable (ΔH r =?8.8kcal/mol,ΔG r (298K)=5.3kcal/mol)than its nitroxyl disul ?de counterpart 6.Unfavorable steric interactions at the Cu center in 6between bulky thiolate tert -butyl and TpCu isopropyl substituents are signi ?cantly relieved by the κ1-N binding mode of t BuSNO in 7.The conversion of 7to the corresponding TpCu(NO)and RSSR ′species proceeds smoothly via the transition state TS7with an activation enthalphy of 5.9kcal/mol (ΔG ?(298K)=19.4kcal/mol)with respect to the starting materials (Scheme 7).

The concerted cleavage of the RS ?NO and Cu ?SR bonds with the simultaneous formation of the RS ?SR bond in the κ1-N ?RSNO adduct iPr TpCu(SR)(R ′SNO)as depicted in TS7accounts for the experimental formation of only unsymmetrical disul ?des.The dispersion energy correction terms calculated with DFT-D3contributes 8?16kcal/mol to the formation enthalpies of 4-7.This clearly suggests the importance of dispersion interactions in stabilizing these intermediates and demonstrates the necessity of the accurate treatment of dispersion e ?ects in these calculations.The reaction of [Cu II ]?SR with RSNO to form [Cu I ](NO)and RS ?SR closes a catalytic cycle for release of NO from S -nitrosothiols provided that copper(II)thiolates [Cu II ]?SR are

formed upon reaction of copper(I)complexes [Cu I ]and RSNOs (Schemes 1and 8).Reaction of iPr2TpCu(NO)(3)with 1equiv C

6F 5SNO at ?40°C in CH 2Cl 2leads to incomplete consumption of 3with formation of iPr2TpCu ?

SC

6F 5(1).Using the steric model iPr TpCu,calculations predict this reaction to be mildly favorable with ΔH

r =+1.9kcal/mol and ΔG r (298K)=?6.7kcal/mol.This reaction is reversible;addition of excess NO to iPr2TpCu ?SC 6F 5at ?40°C forms iPr2TpCu(NO)(3)(Scheme 8).Thus,addition of excess C 6F 5SNO to 1or 3at ?40°C results in the facile Cu-catalyzed

Scheme 5.Energetics of Reactions of Model TpCu ?SMe with MeSNO Calculated with B3LYP-D3/6-311+G(d)

a a Non-dispersion corrected B3LYP enthalpies are given in parentheses.Scheme 6.Energetics of Reactions of Steric Model iPr TpCu ?SC 6F 5with t BuSNO Calculated with B3LYP-D3/6-311G(d)

a

a Non-dispersion corrected B3LYP enthalpies are given in parentheses.Scheme 7.Conversion of iPr TpCu(SR)(N(O)SR ′)to iPr TpCu(NO)and RSSR ′Calculated with B3LYP-D3/6-311G(d)

a

a Non-dispersion corrected B3LYP enthalpies are given in parentheses.Scheme 8.Copper-Catalyzed Release of NO from

RSNOs

decomposition to C6F5S?SC6F5and NO gas over5min, provided that the solution is gently bubbled with N2to remove NO gas as it is formed.Although thermally sensitive around room temperature,this S-nitrosothiol shows little(<5%)decay in the absence of1or3at?40°C in CH2Cl2.

Given the similarity between TpCu?SR models and type1 Cu sites,these studies suggest that RSNOs could directly react with the Cu?SCys moiety at these biological copper centers. The constrained protein environment in which the type1Cu sites are embedded,however,could help resist the loss of a disul?de with formation of a[Cu I](NO)species.Since the SCys moiety is constrained by the protein structure,small molecule RSNOs would be much more freely di?usable than would a product disul?de.Nonetheless,type Cu sites in a number of enzymes such as ceruloplasmin27and ascobate oxidase28have been shown to reversibly react with NO itself. Future reports will describe our e?orts at modeling the nature of the[Cu I](RSNO)intermediates formed upon addition of NO to biologically relevant[Cu II]-SR complexes.

■ASSOCIATED CONTENT

*Supporting Information

Experimental,characterization,and calculational details.This material is available free of charge via the Internet at http:// https://www.wendangku.net/doc/6410127759.html,.

■AUTHOR INFORMATION

Corresponding Authors

nihan.olcum@https://www.wendangku.net/doc/6410127759.html,.tr

thw@https://www.wendangku.net/doc/6410127759.html,

Present Address

?Old Dominion University,Department of Chemistry and Biochemistry,4541Hampton Boulevard,Norfolk,VA23529-0126.

Notes

The authors declare no competing?nancial interest.■ACKNOWLEDGMENTS

T.H.W.thanks NSF(CHE-0957606).N.C.-O.and K.N.H. thank the UCLA Institute for Digital Research and Education (IDRE)and the Extreme Science and Engineering Discovery Environment(XSEDE)for computer time.

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