Fabrizio Badalassi

Enzyme Fingerprints of Activity, and Stereo- and Enantioselectivity from Fluorogenic and Chromogenic Substrate Arrays

A series of stereochemically and structurally diverse fluorogenic and chromogenic substrates for hydrolytic enzymes has been synthesized and used to characterize enzyme activity profiles of esterases, lipases, proteases, peptidases, phosphatases, and epoxide hydrolases. The substrates used are particularly resilient to nonspecific reactions due to their mechanism of activation. The activities recorded with the individual substrates are therefore remarkably reproducible, and enable us to use the overall pattern of activity as a specific fingerprint for the enzyme sample. Fingerprints of activity, and enantio- and stereoselectivity are displayed as arrays of color-scale squares that are easily analyzed visually. Such fingerprints might be useful for quality control, enzyme discovery, and possibly for addressing the issue of functional convergence in enzymes.

A selective HIV-protease assay based on a chromogenic amino acid

(2S,3S)-2-Amino-3-hydroxy-5-(4-nitrophenoxy)pentanoic acid (5) was prepared stereoselectively as the N-Fmoc, O-(tert-butyl)-protected derivative 5a in eleven steps from ethyl (E)-4-benzyloxypent-2-enoate (6). This protected amino acid was used for the solid-phase peptide synthesis of oligopeptides, which serve as sequence-specific chromogenic protease substrates when used in the presence of NaIO4 and bovine serum albumin. The peptide 1 (KRAVNle5EANleNH2 (Nle=norleucine)) allows detection of HIV-protease activity spectrophotometrically at 405 nm.

Improved stereoselective synthesis of both methyl α- and β-glycosides corresponding to the amino sugar component of the E Ring of calicheamicin γ1I and esperamicin A1

Abstract The monosaccharide component (α and β-anomer) of the E Ring of calicheamicin γ1I and esperamicin A1 has been synthetized by an efficient and improved stereoselective procedure starting from methyl 2-deoxy-α- and β-D-ribopyranoside. The synthetic procedure makes use, in each case, of a cyclic sulphate and of the regioselective ring opening of an intermediate activated aziridine.

Efficient enantioselective synthesis of (R)-2-acetyl-2-hydroxy-5,8-dimethoxy-1,2,3,4-tetrahydronaphthalene, the key intermediate in the synthesis of anthracycline antibiotics☆

Abstract A simple, efficient, enantioselective synthesis of (R)-2-acetyl-2-hydroxy-5,8-dimethoxy-1,2,3,4-tetrahydronaphthalene, the key intermediate in the synthesis of anthracycline antibiotics, is described. The synthetic procedure starts with the Sharpless asymmetric dihydroxylation of 2-acetyl-5,8-dimethoxy-3,4-dihydronaphthalene: the diol obtained is regioselectively transformed into the corresponding chloroacetate which is dehalogenated and saponified to give the desired title compound in four steps with satisfactory yield (52%). No separation step is necessary at any point of the synthetic process. An efficient procedure for the synthesis of the starting enone and the stereoselectivity of the methanolysis of the intermediate chloroacetate are also reported.

Stereo- and regioselectivity of cyclization reactions in conformationally restricted epoxy ketones: evaluation of C- versus O-alkylation process

Abstract The intramolecular addition reaction of metal enolates of ketones to oxiranes has been applied to a series of epoxy ketones derived from cyclohexene oxide. γ-Hydroxy ketones (γ-HKs, C -alkylation products) or hydroxy enol ethers (HEEs, O -alkylation products) are obtained, depending on the nature of the cyclic transition state in each case involved and the application of the Furst–Plattner rule. The formation of HEEs by reaction of the same epoxy ketones under acid conditions is also described. In some cases, regioconvergent or chemoselective processes are conveniently obtained.

Cover Picture: Chem. Eur. J. 14/2002

The cover picture shows a two-dimensional color code that can be used to represent catalytic activity (intensity) and enantio- or stereo-selectivity (green/red balance) for the action of enzymes on individual fluoro- and chromogenic substrates in a 6×5 array. The patterns obtained serve as activity fingerprints for each enzyme. The illustration shows the selectivity fingerprints for (from left to right) Pseudomonas cepacia lipase, Candida antarctica lipase, Electrophorus electricus acetylcholine esterase, and Rhizomucor miehei lipase, together with the structure of the enzymes. Color coding is illustrated by connecting two squares in each fingerprint to the corresponding color in the reference grid (bottom). For more details see the article by J.-L. Reymond et al. on p. 3211 ff.