S. Zameo, B. Vauzeilles, J.-M. Beau
SHORT COMMUNICATION
[4] a) E. H. Cordes, W. P. Jencks, J. Am. Chem. Soc. 1963, 85,
2843; b) J. Hine, J. C. Craig, J. G. Underwood II, F. A. Via, J.
Am. Chem. Soc. 1970, 92, 5194; c) A. Bruylants, E. Feytmants-
De Medecis in The Chemistry of the Carbon–Nitrogen Double
Bond (Ed.: S. Patai), Wiley, London, 1970, pp. 465–504. For a
study in a nonaqueous medium, see: N. Giuseppone, J.-M.
Lehn, Chem. Eur. J. 2006, 12, 1715.
analyzed 30 min later. HEWL (2 equiv.) was added to the remain-
ing M2, and the resulting DCL (M5) was stirred, equilibrated at
room temperature for 24 h and analyzed.
Analytical chromatography was performed with a JASCO LC-1500
system equipped with a Phenomenex LUNA C18 (2) 5 µ reverse-
phase HPLC column (150ϫ4.60 mm), with UV-detection at
322 nm. A binary solvent gradient (solvent A: 0.1% trifluoroacetic
acid in H2O 95%/CH3CN 5%, solvent B: 0.1% trifluoroacetic acid
in CH3CN 95%/H2O 5%) was optimized in order to separate most
of the DCL compounds: A 90%/B 10% to A 70%/B 30% over
20 min, with a flow rate of 0.8 mLmin–1.
[5]
S. Zameo, B. Vauzeilles, J.-M. Beau, Angew. Chem. 2005, 117,
987; Angew. Chem. Int. Ed. 2005, 44, 965.
[6]
In this study, the terms aldehyde and imine are used for any
species in fast equilibrium with the true aldehyde and the true
imine, respectively, including hydrate and hemiaminals. An
NMR study performed at a higher concentration showed, how-
ever, the aldehyde and the imine as the sole detectable species.
In this system, and in contrast with the analysis after re-
duction,[5] the other imines were also slightly amplified.
The eluent contained 0.1% TFA, which resulted in a fast dena-
turation of the enzyme and the release of any bound product
on the column. This precluded any composition analysis biased
by the enzyme.
Supporting Information (see footnote on the first page of this arti-
cle): Data used for the estimation of the dissociation constants.
[7]
[8]
Acknowledgments
We gratefully acknowledge the Conseil Général de lЈEssonnefor the
financial support of this study and GlaxoSmithKline for their
interest.
[9]
S. R. Stone, J. F. Morrison, Biochim. Biophys. Acta 1983, 745,
237.
[10] The molar extinction coefficient of imine A·1 at 322 nm was
estimated as approximately 1.6-fold the molar extinction coeffi-
cient of amine A.
[11] The amplification factor is defined as the ratio of the total
(bound+free) imine concentration in the presence of the target
versus the corresponding concentration in the absence of the
target.
[12] Taking into account the affinity of starting amine A for the
binding site of the lysozyme (around 5 m),[5] slightly lower
estimates can be calculated by numerical resolution, namely
0.13 and 0.10 m.
[13] The observed difference might in theory result from the re-
cognition by the active site of the target of a nonreducible spe-
cies, such as a hemiaminal. As previously mentioned,[6] no
hemiaminal could be detected by NMR, which suggests that
the amplified species is indeed the corresponding imine.
[14] As anticipated,[5] no redistribution of the imines in the presence
of the lysozyme was observed with such a library.
Received: September 30, 2006
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Published Online: November 6, 2006
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Eur. J. Org. Chem. 2006, 5441–5444