Table 2 Relative rates (krel) for the hydrolysis of RNA and nucleoside 2A,3A-
monophosphates in the presence of dinuclear Cu(ii) complex 1
This work was supported by the National Institutes of Health
GM 53579).
(
a
Substrate
k
rel
Substrate
k
rel
ApA
CpC
UpU
GpG
87
7.5
5.0
1.0
2A,3A-cAMP
2A,3A-cCMP
2A,3A-cUMP
2A,3A-cGMP
31
5.2
3.0
1.0
Notes and references
†
Hydrolysis of RNA dimers by 1 was followed by HPLC (Ranin). The
following procedure is typical: 2.85 mL of 1 (2.1 mM) in a buffer solution
was mixed with 0.15 mL of an RNA dimer (2.0 mM) in deionized H O.
a
Calculated from the first-order rate constants (kobs) in ref. 7.
2
Aliquots (300 mL) of the reaction mixture were quenched with 50 mM
EDTA (300 mL). After filtration, the quenched solution (15 mL) was injected
onto a C-18 reversed-phase column and eluted for 10–15 min with 0–15%
MeCN in H O containing 0.1% CF CO H (flow rate = 1.0 mL min ). The
2 3 2
eluent was monitored at lmax of the nucleobase (260 nm for adenine and
9
sequences (e.g. ApA) being among the slowest. Replacement
of either adenine group in ApA by a cytidine group diminished
the rate of 1-catalysed hydrolysis threefold. This level of base
selectivity is unprecedented in a simple metal-based ribo-
nuclease mimic that lacks any appended recognition elements
2
1
uracil, 252 nm for guanine and 268 nm for cytidine) by a Ranin UV detector.
The first-order rate constants (kobs) were obtained as slopes of plots of ln(A
) vs. t, where A and A are the integrations of the areas of the HPLC peaks
for the RNA dimer at t = 0 and t, respectively.
0
/
A
t
0
t
(
such as oligonucleotide strands). Dinuclear complex 1 is also
highly active: at a concentration of 2.0 mM it provides over five
orders of magnitude rate acceleration for the hydrolysis of its
best substrate, ApA.10 The data in Table 1 also show that
mononuclear complex 2 has moderate selectivity for adenine.
Apparently, attachment of a bipyridine–Cu(ii) unit to 2 not only
increases its activity but also amplifies its base selectivity.
1 Reviews: B. N. Trawick, A. T. Daniher and J. K. Bashkin, Chem. Rev.,
1998, 98, 939; M. Komiyama, J. Biochem., 1995, 118, 665; J. R.
Morrow, Adv. Inorg. Biochem., 1994, 9, 41; M. W. Gobel, Angew.
Chem., 1994, 106, 1201; Angew. Chem., Int. Ed. Engl.,1994, 33, 1141;
D. S. Sigman, A. Mazumder and D. M. Perrin, Chem. Rev., 1993, 93,
7
We have suggested that a strong p–p stacking interaction
2
295; J. Chin, Acc. Chem. Res., 1991, 24, 145.
between adenine and the bipyridine–Cu(ii) unit may be the
principal reason for the high selectivity of 1 for adenine in
2
Recent examples: (a) P. Hurst, B. K. Takasaki and J. Chin, J. Am. Chem.
Soc., 1996, 118, 9982; (b) M. J. Young and J. Chin, ibid., 1995, 117,
1-catalyzed hydrolysis of nucleoside 2A,3A-cyclic monophos-
1
0577; (c) B. Linkletter and J. Chin, Angew. Chem., 1995, 107, 529;
phates. This same interaction also appears to be responsible for
the base selectivity shown by 1 in promoting the hydrolysis of
RNA. First, the base selectivities for the dinucleotide and cyclic
monophosphate substrates parallel each other, as can be seen in
Table 2. Second, the high selectivity of 1 for adenine in the
hydrolysis of RNA dimers is insensitive to the position of the
adenine group relative to the phosphate bond to be cleaved since
ApC and CpA have almost identical reactivity (Table 1). These
observations are more consistent with association through a less
specific p–p stacking rather than more directed interactions
such as hydrogen bonding and metal coordination to the
nucleobases as major sources of base selectivity. Face-to-face
stacking provides an interaction that is adaptable to different
nucleobase positions. If hydrogen bonding or metal coordina-
tion from 1 to the nucleobases were important in stabilizing the
interaction, it is probable that different base selectivities
between the RNA dimers and nucleoside 2A,3A-cyclic mono-
phosphates would result due to the different orientations and
flexibilities in the two sets of substrates. Likewise, ApC and
CpA should also show quite different reactivities due to the
different positions of adenine in the two substrates.
Angew. Chem., Int. Ed. Engl., 1995, 34, 472; (d) W. H. Chapman, Jr. and
R. Breslow, J. Am. Chem. Soc., 1995, 117, 5462; (e) M. Yashiro, A.
Ishikubu and M. Komoyama, Chem. Commun., 1997, 83; (f) M.
Yashiro, A. Ishikubu and M. Komiyama, J. Chem. Soc., Chem.
Commun., 1995, 1793; (g) M. Komiyama, N. Takeda, M. Irisawa and
M. Yashiro, in DNA and RNA Cleavers and Chemotherapy of Cancer
and Viral Diseases, ed. B. Meunier, Kluwer Academic Publishers,
Boston, 1996, p. 321; (h) J. K. Bashkin, J. Xie, A. T. Daniher, L. A.
Jenkins and G. C. Yeh, ibid., p. 355; (i) R. Haner, J. Hall, D. Husken and
H. E. Moser, ibid., p. 307; (j) D. Magda, R. A. Miller, M. Wright and J.
Rao, ibid., p. 337; (k) J. R. Morrow and V. M. Shelton, New J. Chem.,
1994, 18, 371; (l) R. Ott and R. Kr a¨ mer, Angew. Chem., Int. Ed., 1998,
37, 1957.
3 C. A. Stein and J. S. Cohen, Cancer Res.,1988, 48, 2659.
E. Uhlmann and A. Peyman, Chem. Rev., 1990, 90, 543.
J. F. Milligan, M. D. Matteucci and J. C. Martin, J. Med. Chem., 1993,
4
5
3
6, 1923.
6
For the techniques currently available for RNA sequencing and structure
mapping see: T. D. Tullus, in Bioorganic Chemistry: Nucleic Acids, ed.
S. M. Hecht, Oxford University Press, New York, 1996, p. 1244.
7 S. Liu, Z. Luo and A. D. Hamilton, Angew. Chem., Int. Ed. Engl., 1997,
36, 2678.
8 S. Liu and A. D. Hamilton, Bioorg. Med. Chem. Lett., 1997, 7, 1779.
9
T.Koike and Y. Inoue, Chem. Lett., 1972, 569.
In conclusion, dinuclear Cu(ii) complex 1 has been shown to
function as a highly active artificial ribonuclease. In addition, a
remarkable selectivity among the different nucleotide bases is
seen with particularly effective cleavage of adenine-containing
substrates. We are currently extending these dinuclear Cu(ii)
complex designs to enhance the level of activity and to alter the
base selectivity.
2
10
1
0 The background rate of hydrolysis of ApA at pH 7.5 is ca. 1.6 3 10
2
1
s
, see Y. Matsumoto and M. Komiyama, J. Chem. Soc., Chem.
Commun., 1990, 1050; K. Yoshinari and M. Komiyama, Chem. Lett.,
990, 519.
1
Communication 8/08195F
588
Chem. Commun., 1999, 587–588