S-nitrosothiol and nitric oxide reactivity at zinc thiolates

Article abstract of DOI:10.1021/ic900664r

S-Nitrosothiols undergo reversible transnitrosation reactions at tris (pyrazolyl)boratozinc thiolates ^iPr2TpZn-SR. These zinc thiolates are unreactive toward anaerobic NO but rapidly react with NO in the presence of O₂ or anaerobical

Full text of DOI:10.1021/ic900664r

Inorg. Chem. 2009, 48, 5605–5607 5605
DOI: 10.1021/ic900664r
S-Nitrosothiol and Nitric Oxide Reactivity at Zinc Thiolates
Matthew S. Varonka and Timothy H. Warren*
Department of Chemistry, Georgetown University, Box 571227, Washington, D.C. 20057
Received April 5, 2009
S-Nitrosothiols undergo reversible transnitrosation reactions at tris
(pyrazolyl)boratozinc thiolates iPr2TpZn-SR. These zinc thiolates
are unreactive toward anaerobic NO but rapidly react with NO in
the presence of O2 or anaerobically with NO2 to release the
S-nitrosothiol RSNO with formation of the corresponding zinc nitrate.
domains in metallothioneins (MTs) with concomitant for-
mation of RSNOs and/or disulfides.11 The release of zinc
from MTs by RSNOs and NO has spurred investigation into
the significance of both zinc and RSNOs in cellular signal
transduction11,12 as well as their roles in respiratory func-
tion.13 Similarly, RSNOs and NO reversibly inhibit DNA
transcription of some zinc fingers,14 metalloproteins in which
Zn-thiolate bonds play especially important structural roles.
Matrix metalloproteinases (MMPs) are a class of zinc
enzymes involved in tissue remodeling connected to both
normal and pathological processes such as inflammation,
wound healing, and cancer.15 The His3Zn2+ site responsible
for MMP activity requires prior disruption of a Zn-SCys
linkage in the enzyme’s latent form to allow for substrate
binding and its subsequent cleavage. Both NO and RSNOs
have been proposed to activate this “cysteine switch”, sug-
gesting a molecular basis for NO and RSNO regulation of
MMP activity.16
Nitric oxide (NO) is implicated in numerous biological
roles ranging from vasodilation in the cardiovascular system1
to signaling in the respiratory system2 to host defense against
microbial pathogens.3 While the various NO synthases gen-
erate NO in vivo, NO itself is unstable in the plasma with an
estimated half-life of 3-5 s.4 Considerably more oxygen-
stable S-nitrosothiols (RSNOs) such asS-nitrosocysteineand
S-nitrosoglutathione circulate at near micromolar levels in
the blood.5 Capable of serving as NO and NO+ donors,6,7
RSNOs have been implicated in a wide variety of physio-
logical functions that often mirror those observed for NO
itself.1,8 The nature of the molecular species involved in the
biological reactivity of RSNOs, however, is clouded by the
facile decomposition of RSNOs into free NO and disulfides
by a copper-catalyzed process.6,9
While transnitrosation between RSNOs and free thiols as
well as the corresponding thiolate anions has been observed
in a variety of media,17,18 NO does not react readily under
anaerobic conditions with free thiols HSR or thiolate anions
-SR in the absence of an oxidant.6 In contrast, few molecular
level details are known for transnitrosation at transition-
metal thiolates,19 though reductive nitrosylation of M-SR
The cleavage or formation of zinc thiolate linkages in
biology is often connected to physiological function.10 In
this context, both NO and RSNOs have been reported to
modify Zn-SR linkages in biological environments. For
instance, Maret and co-workers have shown that NO and
RSNOs release Zn2+ ions from the sulfur-rich binding
(11) Chen, Y.; Irie, Y.; Keung, W. M.; Maret, W. Biochemistry 2002, 41,
8360–8367.
(12) Martinez-Ruiz, A.; Lamas, S. Cardiovasc. Res. 2004, 62, 43–52.
(13) Bernal, P. J.; Leelavanichkul, K.; Bauer, E.; Cao, R.; Wilson, A.;
Wasserloos, K. J.; Watkinds, S. C.; Pitt, B. R.; St. Croix, C. M. Circ. Res.
2008, 102, 1575–1583.
*To whom correspondence should be addressed. E-mail: thw@
georgetown.edu.
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r
2009 American Chemical Society
Published on Web 05/26/2009
pubs.acs.org/IC

5606 Inorganic Chemistry, Vol. 48, No. 13, 2009
Scheme 1. Transnitrosation (Top) and Reductive Nitrosylation (Bot-
tom) at Metal Thiolates [M]-SR
Scheme 3. Transnitrosation Equilibria at Zinc Tholates in CDCl3
Scheme 2. Synthesis of Zinc Thiolates 2-5
of each reactant, transnitrosation between BnSNO with
the bulkier iPr2TpZn-SBut complex is qualitatively slower
than with the smaller cyclohexyl derivative iPr2TpZn-SCy. To
more quantitatively define the time scale for transnitrosation,
we monitored degenerate transnitrosation between iPr2TpZn-
SBn and BnSNO by 1H NMR spectroscopy in CDCl3
(Scheme 3).
linkages with NO is possible at redox-active metal centers
(Scheme 1).20 Such transnitrosation reactions are thought to
proceed via nitroxyl disulfide intermediates [RSN(O)NSR0]-
theoretically considered by Houk et al.21 and observed via
15N NMR spectroscopy by Estrin et al. in the exchange
between S-nitrosocysteine ethyl ester and its thiolate anion in
methanol.18
Irradiation of the S-nitrosothiol PhCH2SNO resonance
decreased the intensity of the iPr2TpZn-SCH2Ph signal,
demonstrating saturation transfer between these two exchan-
ging species. The pseudo-first-order rate constant via satura-
tion transfer (Figure S5 in the Supporting Information)
increases linearly with an increase in the RSNO concentra-
tion, leading to a second-order rate law k[Zn-SR][RSNO]
with a rate constant 2.0(2) M-1 s-1 at 60 °C. No significant
decomposition (less than 5%) of the RSNOs was observed
under these conditions. Transnitrosation with RSNOs at zinc
thiolates is significantly faster than alkylation26 or disulfide
exchange23 in related model complexes. For instance, trans-
nitrosation of iPr2TpZnSCH2Ph (2) with PhCH2SNO is
500 times faster than alkylation by MeI in CDCl3 at 60 °C
[k = 3.8(3) ꢀ 10-3 M-1 s-1; Scheme 4].
We recently demonstrated transnitrosation between the
β-diketiminatozinc thiolate {[Me2NN]Zn}2(μ-SBut)2 and Cy-
SNO to give equilibrium quantities of {[Me2NN]Zn}2-
(μ-SCy)2 and tBuSNO.19 Unfortunately, the dinuclear nature
of these zinc complexes clouded the thermodynamic prefer-
ences for transnitrosation equilibria. Herein we utilize well-
defined tris(pyrazolyl)boratozinc complexes22 iPr2TpM-SR
to examine RSNO and NO reactivity at strictly mononuclear
zinc thiolate sites.
Tris(pyrazolyl)boratozinc thiolates23-25 iPr2TpZn-SR [R =
None of the zinc thiolates 2-5 react with NOgas in CDCl3
under anaerobic conditions; standing for 12 h with ca. 8 equiv
t
Bn (2) Cy (3), Bu (4), C6F5 (5)25] are prepared by salt
metathesis reactions between iPr2TpZn-Cl (1) and the corre-
sponding thallium thiolate Tl-SR (Scheme 2). Reaction of the
secondary and tertiary zinc thiolates 3 and 4 with BnSNO
(generated by the addition of Tl-SBn in CDCl3 to dry NO
[BF4]) allows for transnitrosation equilibria to be observed at
room temperature in CDCl3, as outlined in Scheme 3. While
there is no significant bias in the equilibrium between the
secondary zinc thiolate 3 and the primary S-nitrosothiol
BnSNO [Keq = 0.65(8)], transnitrosation favors the forma-
tion of the primary zinc thiolate 2 and tertiary S-nitrosothiol
tBuSNO [Keq = 59(5)]. These equilibria data qualitatively
mirror the trend observed in reactions between zinc thiolates 3
and 4 and the thiol HSBn [Keq = 3.2(4) and 750(50),
respectively]. Moreover, the reaction between tertiary zinc
thiolate 4 and the electron-poor S-nitrosothiol C6F5SNO
results in complete conversion to the zinc arylthiolate 5 and
tBuSNO.
1
of NO results in no change in their H NMR spectra. No
reaction is observed with excess O2 under similar conditions.
The addition of 8 equiv of NO to a O2-saturated CDCl3
solution of 4, however, results in an immediate color change
to green owing to essentially the quantitative formation
of tBuSNO confirmed by its 1H NMR signal at δ 1.92 ppm
(Scheme 5). The addition of 8 equiv of anaerobic NO2 gas to
a CDCl3 solution of 4 also provides similar results.
In the reaction of 4 with NO/O2 or NO2, only one new
iPr2TpZn species 7 is produced. X-ray analysis of the
colorless product identifies it as iPr2TpZn(NO3), which
has unsymmetric κ2 bonding of the nitrate anion to the
˚
Zn center with Zn-O distances of 1.954(2) and 2.488(3) A.
This unsymmetric bonding mode is seen in other crystal-
lographically characterized TpZn(NO3) complexes.27
Because N2O3 has been suggested as a potential nitrosat-
ing reagent in reactions of NO under aerobic conditions,
which could generate the nitrite anion after transfer of
NO+,28 iPr2TpZn(NO2) (8) was independently prepared
While the above transnitrosation equilibria were generally
complete within 5 min at approximate 0.1 mM concentrations
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Communication
Inorganic Chemistry, Vol. 48, No. 13, 2009 5607
Figure 1. X-ray structures of iPr2TpZn-SBut (4), iPr2TpZn(NO3) (7), and iPr2TpZn(BF4) (9) (4 was collected at 173 K, and 7 and 9 were collected at 100 K).
Scheme 4. Alkylation of 2 with MeI to Give Zinc Iodide 6
Scheme 6. Independent Synthesis of Zinc Nitrate 7 and Zinc Nitrite 8
Scheme 5. NO and NO2 Reactivity of Zinc Thiolate 4
Scheme 7. Nitrosation of Zinc Thiolate 4 with NOBF4
binding of the BF4 anion has been observed in Ni30 and Cu31
complexes.
These studies with well-defined, mononuclear zinc thio-
lates demonstrate that thiolate nitrosation with reagents
capable of serving as equivalents of NO+ such as RSNOs
and aerobic NO is quite facile, particularly so when consider-
ing the steric bulk of the iPr2Tp ancillary ligand. Owing to
the redox stability of the Zn2+ ion, the Zn-thiolate
bond exhibits no anaerobic NO reactivity under similar
conditions. Moreover, transnitrosation with the S-nitro-
sothiol PhCH2SNO is much faster than the conceptually
related alkylation reaction with MeI. Future studies will
explore transnitrosation at various zinc coordination envir-
onments as well as probe the nature of the zinc intermediates
in transnitrosation.
by the reaction of Zn(ClO4)2 with iPr2TpK and NaNO2 in
MeOH (Scheme 6). The X-ray structure of the zinc nitrite
complex 8 is closely related to that of 7 with Zn-O
˚
distances of 1.981(2) and 2.430(2) A (Figure S14 in the
Supporting Information). Importantly, nitrate/nitrite
exchange between iPr2TpZn fragments is slow in a CDCl3
solution at room temperature; an overlapping eight-line
pattern is observed in the 1H NMR spectrum of a mixture
of 7 and 8, resulting from four sets of diastereotopic
CHMe2 groups. Thus, our evidence points to the sole
formation of the zinc nitrate 7 when zinc thiolate 4 is
exposed to NO with an excess of O2. Under these condi-
tions, the active nitrosating reagent behaves analogously
to NO2 which can exhibit electrophilic reactivity owing to
its equilibrium with N2O4 reacting as [NO+][NO3-].29
Similarly, treatment of 4 with NOBF4 results in the forma-
tion of iPr2TpZn(BF4) (9) along with tBuSNO (Scheme 7). The
zinc-containing product 9 was confirmed by an independent
synthesis through the salt metathesis reaction of 1 with
AgBF4. The BF4 anion exhibits κ2 coordination in 9 with
Zn-F1 and Zn-F2 bond distances of 1.997(2) and 2.379(2)
Acknowledgment. T.H.W. thanks the NSF (CAREER
program) and ACS PRF (43048-AC3) for funding. M.S.
V. is grateful to the Georgetown University Chemistry
Department for an Espenscheid Fellowship.
Supporting Information Available: Complete experimental
details as well as X-ray crystallographic data (CIF). This
material is available free of charge via the Internet at
http://pubs.acs.org.
˚
A, respectively. The X-ray structure of 9 appears to be an
unusual example of Zn-BF4 coordination, though related κ2
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