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Binuclear Metal Complexes as Efficient Intermediaries in Biochemically Relevant Hydrolysis Reactions.

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HIGHLIGHTS
Binuclear Metal Complexes as Efficient Intermediaries
in Biochemically Relevant Hydrolysis Reactions
Michael W. Gobel"
Enzymes set chemists a high standard for synthetic catalysts
with their often astoundingly high selectivity and simultaneous
enormous acceleration of reactions. This goal is very difficult to
achieve, let alone to better. The elucidation of enzyme mechanisms is therefore counted among the important tasks of science,
to which a wide variety of research disciplines can contribute. In
recent years X-ray crystal structure analysis, in particular, has
inade available a wealth of detailed structural information that
can serve a foundation for the formulation of mechanistic proposals. Further refinement of such models is then possible
through site-directed mutagenesis and calculations. In the light
of the success of scientific methods, which in a certain sense can
give a multifaceted view of the problem, the role preparative
chemistry may play nowadays in elucidating enzyme catalysis is
questioned. Is the synthesis of low molecular weight models of
enzymes actually still relevant?
I am convinced that this question can be answered with a
decisive yes. One misunderstanding, however. must be avoided.
Models of enzymes that describe the complex structure and
dynamics of a proteinogenic catalyst are, as a rule, not achieved
in the attempts of synthetic chemists. This is also not the goal of
such research. The aim of investigations on small, well-defined
molecules is Far more to probe a particular aspect of the enzyme
in question, for example. to consider in isolation the special
arrangement of functional groups obtained from crystal structure data and to test the relevance of such an arrangement with
respect to the reaction mechanism. In this way individual principles of function are determined, which are useful for a deeper
understanding. Another advantage of the reductionistic view
lies in the potential for using the principles found in the model
to construct totally novel. enzyme-analogous catalysts that
could attain practical importance in the future.
A good example for the value of such model systems is the
series of papers that have recently appeared on binuclear metal
complexes. Beside the long-known iron-sulfur proteins of the
respiratory chain. enzymes and proteins have been described
recently in which the active sites contain two or three interacting
metal ions (for example, Zn, Fe. Cu).['] The biological tasks
cover oxygen transport to oxygenase and hydrolase activity.
Since the structure-function relationship of these enzymes are
by no means solved, the synthesis of simplified model complexes
[*] Dr. M . W. Gobel
Institut fur Orgsnische Chemie der Universitiit
Marie-Curie-Strnsse 11. D-60439 Frankfurt am Main
Telefax: In( code + (6Y)5800-9250
is an important goal. Typically. symmetrical ligands linked by
meta-substituted arene units are used. In this way, with 1 Karlin
et al. have prepared a binuclear copper complex along the lines
of hemocyanin, the copper-containing 0,-transport protein of
molluscs. At low temperature 1 binds 0, reversibly.[*"' The oxygen associate of complex 2 is more stable.[2b1In the presence of
dimethylformamide ( D M F ) or dimethylacetamide. however,
this associate reacts instantaneously to form 3, whose structure
2
fi
O2 associate of 2
3
has been solved by X-ray crystallography. In analogous copper
complexes the remarkable redox reaction of hydroxylation of
the arene and oxidation of the copper ions occurred earlier.[2c1
Completely unexpected was the fate of the amide: it was hydrolyzed to the formate ion. In the light of the high kinetic
stability of cdrbOXyk amides, whose half-lives in neutral water
lies in the range of years, the hydrolysis of D M F within seconds
is a new speed record! Where can the reason for this enormous
acceleration be found? The presence of both copper ions appears to be essential. While 4 hydrolyzes D M F with a rate constant of about 0.3 h - ', no reaction can be detected with the
mononuclear complex 5.[3'Probably the first C u ion of 4 coordinates a hydroxide ion, while the second Cu cation binds the
HIGHLIGHTS
4
5
oxygen atom of DMF and activates it as Lewis acid for the
intramolecular addition of the nucleophile (Scheme 1 ) . With
respect to its reactivity. 4 is
comparable with the known
metal catalysts of amide hydrolysis;[31nevertheless, the
oxygen associate of 2 is unparalleled. As is known, the
nucleophilicity of OOHions is drastically enhanced
over that of the OH- ion on
account of the x effect. The
NR2
Scheme 1. postulated mechanisnl "1.
authors therefore surmise
the amide hydrolysis uith 4 3s catalyst.
that the species that is responsible for the hydrolysis
is a copper complex with peroxido ligand that arises in the
primary redox reaction. Scheme 2 depicts the assumed mechanism. The study of complex 2 thus led not only to the answer of
the original question on the
binding of oxygen in copper
complexes, but simultaneously to a novel approach
to the construction of artificia1 peptidases.
In nature multinuclear
metalloenzymes,
under
which the RNAse H of the
HIV reverse-transcriptase is
Scheme 2. Postulated mechanism of
classified, Serve as catalysts
the amide hydrolysis \rith 2 as catalyqt.
for several acyl and phosphoryl transfer reactions.
Chin et al. chose a binuclear copper complex related to 1 to
accelerate the reaction of a phosphoric acid diester anion
(Scheme 3).14] This reaction can be considered a model for the
first step of the hydrolysis of RNA, in which the alcohol function
of the side chain attacks the phosphoryl center as nucleophile in
a ring closure reaction. As in the case already described. the
binuclear copper complex was compared with the analogous
mononuclear complex: the approximately 50 times larger rate
constant of the binuclear complex shows that here, too, the two
metal ions probably ~ o o p e r a t e . 'Because
~]
on cyclization of the
substrate the transition state structures have a higher negative
charge density at the phosphoryl group, the interaction with
several electrophilic metal sites is advantageous. Analogous zinc
complexes have already been used for the hydrolysis of similar
model substrates."] The ultimate test for every synthetic hydrolase, however, is the phosphoric diester group of DNA. Since an
entropically favored intramolecular course of reaction is not
Scheme 3. Model reaction
for the first step of the hydrolysis of RNA B =
base.
possible, in contrast to RNA, because of the missing hydroxyl
function at C2' in DNA, these phosphoric acid esters show an
extreme kinetic stability to substitution. With the redox-active
Ce3'i0, system. Chin et al. have now been able to perform
the h-vdrolytic cleavage of deoxyribodinucleotides as well in
convincing style."] Although the mechanism of the hydrolysis
has not been completely elucidated, the assumption of a
reaction of Ce3+ ions with oxygen to give a complex of two
Ce4' ions and the dianion 0;- as active agent (Scheme 4) seems
plausible. The pathway should be characterized by a similar
combination of electrophilic and nucleophilic properties as
the intermediate in the amide hydrolysis observed by
Karlin et al.
Hd
Hd
Scheme 4. Conjectured mechanism of the hydrolysis of deoxyribodinucleotides elfected by Ce'+:O,
To achieve an electrophilic activation (similar to a Lewis acid)
of substrates, enzymes are not, however, exclusively dependent
on metal ions. Often guanidinium ions of arginine side chains
also perform this function. In this way the Staphylokokkus
nuclease accelerates the hydrolysis of phosphoric acid diester
anions by sixteen orders of magnitude (!), although the active
site contains only one calcium ion. The decisive factor is the
additional presence of two guanidinium ions that display both
the electrostatic and the structural complementarity of the dianionic, trigonal-bipyramidal transition structures of the substrate
hydrolysis. To date already three research groups have attempt-
HIGHLIGHTS
ed to employ the bis(guanidinium) substructure of the Staphylokokkus nuclease as active principle for synthetic catalysts with
phosphodiesterase activity."] In compounds 6 and 7 the cationic
groups are linked to each other in such a way that four hydrogen
COOC2H5
6
7
bonds can form simultaneously to a tetrahedral or trigonalbipyramidal 0x0 anion to be complexed. As was hoped, 7 and
analogous compounds strongly accelerate substitution reactions of phosphoric acid diesters in dipolar, aprotic solvents.[8c.d1Anslyn, Smith et al. could even catalyze the cleavage
of R N A in water with 6.[8b1
The binuclear Co complex 8 described by Czarnik et al.[91
differs from those already mentioned. The two metal ions are
held so far apart by the
spacer that no direct
dimerization of the
two subunits through
oxygen is possible.
Complex 8 accelerates
the hydrolysis of phosphoric acid diester anions by seven orders of
magnitude, but still
does not exceed the ac8
tivity of mononuclear
complexes. In co3+
complexes nucleophile and phosphoric acid diester bind to the
same metal ion and react intramolecularly via a four-membered
transition state (Scheme 5. left). The high efficiency of 8 is therefore not attributable to a cooperativity of the two Co3+ions. On
the other hand 8 increases the rate of hydrolysis of phosphoric
acid monoesters about ten times more than the corresponding
mononuclear complexes. Probably one metal ion coordinates
the nucleophilic water molecule, while the second binds the
phosphoryl group in the form of a four-membered ring.[31The
Afi,qiw. ( ' / i c m .
/fir.
Ed. D i g / . 1994. 33. N o . 11
reaction of the phosphoric acid monoester can therefore profit
from the special arrangement of the metal ions in 8 (Scheme 5,
right).
This small selection of examples is intended to demonstrate
that synthetic metal complexes provide a plethora of subtle insights into the structure-activity relationships, which may also
be valid for enzymes. Finely adjusted modifications are much
more difficult to achieve in proteins and still more difficult to
test. Moreover, the presented model systems show substantial
reactivities in spite of their simple construction. The compounds
are indeed of practical interest, perhaps as artificial proteases or
as synthetic ribonucleases that---together with antisense oligonucleotides-can effect a hydrolytic deactivation of pathogenic
messenger-RNA.
/
/
Scheme 5. Postulated mechanism for the hydrolysis of phosphoric acid esters.
German version Angrit Chem 1994, 106, 1201
111 a) K. D. Karlin, Science 1993, 261, 701 -708: b) B. L. Vallee, D. S. Auld. Brochemistrj 1993, 32, 6493 -6500.
[ 2 ] a) P. P. Paul. Z. Tyeklir, R. R. Jacobson, K. D. Karlin. J. Am. CIi~rii.S O .1991,
113, 5322-5332: b) N. N. Murthy. M. Mahroof-Tahir, K . D. Karlin. ihid. 1993.
115, 10404-10405: c) M. S. Nasir, B. 1. Cohen. K. D. Karlin, ihid. IY92. 114,
2482-2494.
[ 3 ] The hydrolysis of amides is. however, also possible at mononuclear metal complexes: J. Chin, Acc. Chem. Rm. 1991. 24, 145-152.
[4] M. Wall, R. C. Hynes, J. Chin, Angrw. Chrm. 1993, 105. 1696 -1697. Anpew.
CIwm. hi.Ed. Engl. 1993, 32, 1633-1635.
[S] The authors point out, however, that the different kinetics could alao he related
to different catalyst -substrate affinities.
(61 a ) Binucledr Zn complex as model for the active site of the phospholipdse C: S.
Uhlenbrock. B. Krebs, Arigew. Chrni. 1992. 104, 1631 --1632; Angtw Chern. hi,
Ed. Engi. 1992, 31, 1 6 4 7 ~1648: b) hydrolysis of phosphoric acid esters at binuclear Zn complexes: S . Hikichi, M . Tanaka. Y Moro-oka, N. Kitajima. J. (%em.
Soc. Cliein. Commwr. 1992. 814-815.
[7] B. K. Takasaki. J. Chin. J. Am Chrni. Soc. 1994, I /6,1121 11 22.
[8] a) D. M. Kneeland, K. Ariga. V. M. Lynch, C -Y. Huang, E. V. Anslyn, ,I Am.
Clirm. Soc. 1993, 115. 10042- 10055. b) J. Smith. K. Ariga. E. V. Anslyn. h d .
1993, 115. 362 364: c) V. Jubian. R. P. Dixon. A. D. Hamilton, !hid. 1992. 114,
1120- 1121: d) R. GroD, G. Diirncr, M. W. Gobel, Lirhig.\ Ann. ('hem. 1994.
49 -58.
[9] D. H. Vance. A. W. CLarnik, J. A m . C h m . S o ( . 1993. 115. 12 165 12 166.
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