Rapid hydrolysis of 2',3'-cAMP with a Cu(II) complex: Effect of intramolecular hydrogen bonding on the basicity and reactivity of a metal- bound hydroxide

Full text of DOI:10.1021/ja981227l

4710
J. Am. Chem. Soc. 1999, 121, 4710-4711
Rapid Hydrolysis of 2′,3′-cAMP with a Cu(II)
Complex: Effect of Intramolecular Hydrogen
Bonding on the Basicity and Reactivity of a
Metal-Bound Hydroxide
Mark Wall, Barry Linkletter, Dan Williams,
Anne-Marie Lebuis, Rosemary C. Hynes, and Jik Chin*
Department of Chemistry, McGill UniVersity
Montreal, Canada H3A 2K6
ReceiVed April 13, 1998
ReVised Manuscript ReceiVed December 14, 1998
In general it is difficult to study the hydrolysis of biologically
important phosphate diesters with poor leaving groups (e.g., DNA)
because of their enormous stability. Consequently, activated
phosphate diesters with good leaving groups (e.g., bis(2,4-
dinitrophenyl)-phosphate (BDNPP), bis(p-nitrophenyl)phosphate
Figure 1. HPLC traces of 1 (1 mM) promoted cleavage of 2′,3′-cAMP
(0.05 mM) at 25 °C, pH 5.0. Retention times are as follows: 3′-AMP )
3.5 min; 2′,3′-cAMP ) 7.5 min; 2′-AMP ) 8 min. Reaction times are
from foreground to background, 6, 30, 120, 306, and 630 s.
(
BNPP), 2-hydroxy-propyl p-nitrophenyl phosphate (HPNP)) are
Table 1. Apparent Second-Order Rate Constants (M- s-1) for
Hydrolysis of Phosphate Diesters at 25 °C
1
often used as substitutes for testing the reactivity of newly
developed artificial phosphodiesterases. However, the mechanism
for catalytic hydrolysis of phosphate diesters with good leaving
is not always the same as that for those with poor leaving groups.1
To this extent, 2′,3′-cyclic adenosine monophosphate (2′,3′-cAMP)
is an ideal substrate for studying the effects of catalysts on
hydrolyzing phosphate diesters with poor leaving groups since
the cyclic phosphate hydrolyzes almost as rapidly as BDNPP
under basic conditions due in large part to the five-membered
ring strain (Table 1). Here we compare the reactivity of three
Cu(II) complexes (1, 2, and 3) for hydrolyzing 2′,3′-cAMP.2
BDNPP (k) BDNPP (rel) 2′,3′-cAMP (k) 2′,3′-cAMP (rel)
-3
-3
OH 3.2 × 10
1
1.1 × 10
1
4.6
-4
-1
2
-3
-2
3
2
1
5.7(2) × 10
8.0(2) × 10
2.0(1) × 10
1.8 × 10
2.5 × 10
6.3 × 10
5.1(2) × 10
6.2(2) × 10
3.8(1) × 10
-
1
5.6 × 10
3
4
3.5 × 10
metal complex. An interesting dinuclear Cu(II) complex that
provides comparable regiospecificity for hydrolyzing 2′,3′-cAMP
has been recently reported. Surprisingly, 1 is over 100 times more
reactive than even Ln(III) ions or complexes (Table 1).
6
7
a
Potentiometric titration reveals that the pK values of the
protonated metal-hydroxides in 1, 2, and 3 are 5.5, 7.0, and 8.2,
8
respectively. It is remarkable that the amino group in 1 lowers
a
the pK of the metal-bound water molecule given that it is an
electron-donating group. It appears that the amino group is acting
as a hydrogen bond donor to the metal-bound water molecule,
thereby lowering the pK
as a hydrogen bond acceptor, the pK
a
(1H). If the amine nitrogen was acting
of the coordinated water
a
molecule should have increased. To test this, we synthesized an
analogue of 1H with the two hydrogen bond donating amine
groups replaced with two hydrogen bond accepting groups (4H).
2
,9-Diamino-o-phenanthroline was synthesized according to a
3
literature procedure. [(2,9-Diamino-o-phenanthroline)CuCl
prepared from a methanolic solution of CuCl and 2,9-Diamino-
o-phenanthroline. Copper complexes 1, 2, and 3 were freshly
generated in water from the corresponding chlorides and 1 equiv
NaOH. Hydrolysis of 2′,3′-cAMP was monitored by HPLC as
2
] was
a
As anticipated, the pK of the metal-bound water in 4H (7.7) is
2
9
significantly higher than that of 1H (or the conjugate acid of 2).
4
described previously. The reaction solutions were buffered by
the metal complexes.5 A typical HPLC plot for 1-promoted
cleavage of 2′,3′-cAMP (0.05 mM) at pH 5 and 25 °C is shown
in Figure 1. It is evident from the HPLC plot that the cleavage
reaction occurs hydrolytically producing 3′-AMP and 2′-AMP in
a ratio of about 16 to 1. This represents the most regiospecific
hydrolysis of the cyclic phosphate with a simple mononuclear
(1) (a) Jencks, W. P.; Kirsch, J. F. J. Am. Chem. Soc. 1964, 86, 837. (b)
Menger, F. M.; Ladika, M. J. Am. Chem. Soc. 1987, 109, 3145. (c) Bashkin,
J. K.; Jenkins, L. A. J. Chem. Soc., Dalton Trans. 1993, 3631.
To examine the possiblity of the intramolecular hydrogen
bondings in 1H, we determined the crystal structure of the
(
2) For some other studies on hydrolysis of phosphates with Cu(II)
complexes, see: (a) Kovari, E.; Kramer, R. J. Am. Chem. Soc. 1996, 118,
2704. (b) Morrow, J. R.; Trogler, W. C. Inorg. Chem. 1988, 27, 3387. (c)
1
Stern, M. K.; Bashkin, J. K. Inorg. Chim. Acta, 1997, 263, 49. (d) Liu, S.;
Hamilton, A. D. Tetrahedron Lett. 1997, 38, 1107. (e) Burstyn, J. N.; Deal,
K. A. Inorg. Chem. 1993, 32, 3585.
(6) Liu, S.; Luo, Z.; Hamilton, A. D. Angew. Chem., Int. Ed. Engl. 1997,
36, 2678.
(7) Breslow, R.; Huang, D.-L. Proc. Natl. Acad. Sci. U.S.A. 1991, 88, 4080.
(
3) Ogawa, S.; Yamaguchi, T.; Gotoh, N. J. Chem. Soc., Perkin Trans. 1.
a
(8) The pK values of the two amino groups in 1 (or 1H) are expected to
1
974, 976.
a
be much higher than 5.5. For example, the pK of the amino group in the
(
4) Linkletter, B.; Chin, J. Angew. Chem., Int. Ed. Engl. 1995, 34, 472.
Co(III) complex of dipyridylamine has been shown to be about 5. Wong, Y.
(
5) We were not able to find a buffer system that does not decrease the
J.; Petersen, J. D.; Geldard, J. F. Inorg. Chem. 1985, 24, 3352.
activity of 1 for cleaving 2′,3′-cAMP. The pH of the reaction solution did not
change by more than 0.1 unit during the cleavage reaction.
a
(9) The pK of the metal bound water in the Cu(II) complex of 5,5′-
diaminobipyridyl (5.65) is comparable to that of 1H.
1
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Communications to the Editor
J. Am. Chem. Soc., Vol. 121, No. 19, 1999 4711
2
′,3′c-AMP may be because the amino group hydrogen bonds to
13
the OH group in 5 (as in 1H). This hydrogen bonding should
acidify the OH group, making it easier for this group to
deprotonate and thus facilitate the expulsion of the leaving group.
This deprotonation may well be coupled to solvent-mediated
protonation of the leaving group oxygen. The extent of this proton
switch at the transition state is expected to be greater for phosphate
diesters with poorer leaving groups.14 Hence the difference in the
reactivity of 1 and 2 for hydrolyzing BDNPP is only about 20-
fold while that for 2′,3′-cAMP is about 600-fold (Table 1). The
a
lowering of the pK in 1H is expected to lower the nucleophilicity
of the metal-hydroxide. However, this appears to be more than
compensated for by the subsequent ease in the deprotonation of
the OH group in 5.
Figure 2. Crystal structures (ORTEP representation; ellipsoids at 50%
probability level) of the dichloro complex of 1 (A) and 2a (B). The two
phenyl groups of bathocuproine in 2a have been omitted for comparison.
dichloro complex of 1 (Figure 2A).10 The range of distances
required between a nitrogen donor and a chlorine acceptor for a
hydrogen bond in the solid state is 3.10-3.26 Å11 and both N-Cl
distances in Figure 2 are well within this range (3.13 and 3.17
Å). For comparison, a crystal structure of a Cu(II) complex where
the amino groups have been replaced by methyl groups has been
obtained. In the copper chloride complex of bathocuproine (2a,
Figure 2B)12 there is no possibility of hydrogen bonding inter-
actions and the distances between the chlorine atoms and the
methyl carbon atoms are considerably longer (3.62 and 3.53 Å,
respectively).
It is unlikely that the amino group is acting as a general base
catalyst to directly deprotonate the OH group in 5 since its basicity
is expected to be too weak (For example, the pK value of the
a
15
conjugate acid of 2-aminopyridinium is only -7.6. ). It is also
unlikely that the amino group is acting as a nucleophilic catalyst
for hydrolyzing 2′,3′-cAMP since 6 does not hydrolyze to any
appreciable extent under the same experimental conditions used
to hydrolyze 2′,3′-cAMP to completion.
The reactivities of the free hyroxide and the three metal-
hydroxide complexes for hydrolyzing BDNPP and 2′,3′-cAMP
a
are listed in Table 1. At solution pH below the pK of the
4
coordinated water molecules, complex 1 is about 2 × 10 times
6
more reactive than 2 and about 4 × 10 times more reactive than
3
a
for hydrolyzing 2′,3′-cAMP. Above the pK of the coordinated
2
water molecules, complex 1 is over 6 × 10 times more reactive
3
than 2 and over 7 × 10 times more reactive than 3 (Table 1).
The greater reactivity of 1 cannot be due to an increase in the
equilibrium constant for complexation of the phosphate diester
since dimethyl phosphate does not significantly inhibit the reaction
up to 0.1 M. The simple mononuclear Cu(II) complex (1) is about
9
In conclusion, 1 provides over (1 × 10 )-fold rate acceleration
1
00-fold more reactive than the recently reported dinuclear
6
over the background hydroxide rate (1 mM metal complex at pH
6
Cu(II) complex for hydrolyzing the cyclic phosphate.
, 25 °C) for hydrolyzing 2′,3′-cAMP by a novel mechanism.
It is well-known that cis-aquahydroxy metal complexes dimer-
ize and that the dimers are not reactive for hydrolyzing
Complex 1 is by far the most reactive transition-metal complex
reported to date for hydrolyzing 2′,3′-cAMP.16
phosphates.2e,4 The rate of hydrolysis of 2′,3′-cAMP increases
linearly with increase in concentration of 1 or 2, indicating that
the metal complexes do not dimerize significantly under our
Acknowledgment. We thank NSERC, Pioneer Hi-Bred International
Inc., and the U.S. Army research office for support of this work. We
also thank Dr. Kimin Park (at Pohang Institute of Technology, Korea)
for his kind help with Figure 2.
4
experimental conditions. Potentiometric titrations of 1, 2, and 4
at various concentrations also reveal that the metal complexes
do not dimerize.
We propose that the mechanism for 1- and 2-promoted
hydrolysis of 2′,3′-cAMP involves intramolecular nucleophilic
attack by the metal-hydroxide on the coordinated phosphate
JA981227L
(
13) Consistent with this interpretation, compound 4H is not active for
hydrolyzing 2′,3′-cAMP under our experiemental conditions up to pH 8.
14) Extent of bond making and breaking in transition state 5 is not
2
diester. The reason 1 is more reactive than 2 for hydrolyzing
(
(
10) Crystal structural data for brown thin plates of 1: monoclinic, space
indicated. This reaction may well be concerted (Hengge, A. C.; Cleland, W.
W. J. Am. Chem. Soc. 1990, 112, 7421) since the metal-hydroxide is expected
to be as good a leaving group as p-nitrophenol. Unfortunately, kinetic solvent
group P-1, a ) 6.9839(13) Å, b ) 10.4179(14) Å, c ) 18.806(3) Å, R )
8
4.683(11)°, â ) 80.406(13)°, γ ) 77.085(12)°, Z ) 4; R ) 0.1000, GOF )
.095. Crystal structural data for red crystals of 2b: monoclinic, space
group P-1, a ) 14.300(3) Å, b ) 15.759(3) Å, c ) 11.8256(15) Å, R )
00.423(13)°, â ) 102.141(12)°, γ ) 107.165(13)°, Z ) 4; R ) 0.069, GOF
3.30.
11) Macgillavry, C. H., Rieck, G. D., Eds. International Tables for X-Ray
Crystallography; The Kynoch Press: Birmingham, 1962; vol 3.
12) Linkletter, B. PhD dissertation, Hydrolytic CleaVage of RNA by Metal
Complexes; Department of Chemistry, McGill University, Canada, 1995.
1
isotope effect could not be measured because 1 is not soluble in D
to 0.1 mM, whereas it is soluble in H O up to 2 mM. It may be that the
intramolecular hydrogen bond in 1H is weakened in D O.
(15) Jencks, W. P.; Regenstein, J. Handbook of Biochemistry; Chemical
Rubber Company: Cleveland, OH, 1970; pp J-150.
(16) Ligand free Ce(IV) is more reactive than 1. See, for example: (a)
Takasaki, B. K.; Chin, J. J. Am. Chem. Soc. 1994, 116, 1121. (b) Sumaoka,
J.; Miyama, S.; Komiyama, M. J. Chem. Soc., Chem. Commun. 1994, 1755.
2
O down
2
1
)
2
(
(