David B. Berkowitz

(α-Monofluoroalkyl)phosphonates: a class of isoacidic and “tunable” mimics of biological phosphates

Abstract In the early 1980s, Blackburn and McKenna suggested that α-fluorination might lead to phosphonates that better mimic natural phosphates. Although α-monofluorination produces phosphonates with “matching” second pKa values, the α,α-difluorinated phosphonates have received more attention in the past decade or so. Recently, reported enzyme kinetic data on the α-monofluorinated phosphonates from the O’Hagan lab and from our lab suggest that the CHF stereochemistry does affect enzyme-binding, thereby providing an additional variable that may be tuned to achieve optimal binding to an active site of interest. This asymmetry also appears in structural data from the groups of Barford/Burke and Tracey on PTP1B complexes with bound α,α-difluorinated phosphonate inhibitors. In those complexes, only one of two prochiral fluorine atoms appears to interact appreciably with the enzyme. Namely, it is thought that the pro-R (Fsi) fluorine is engaged in an important hydrogen bond with the Phe-182 amide NH. Available methods for the synthesis of this class of α-monofluorinated phosphonates are reviewed. A new convergent approach, developed at Nebraska, in which the potassium anion of (α-fluoro-α-phenylsulfonylmethyl)phosphonate is used to displace primary triflates is also described. This method is particularly convenient as it allows one to perform a “fluorinated phosphonate scan” of an active site of interest (in what follows, we use this expression to designate the synthesis and evaluation of a complete set of the CH2–, CF2– and both stereoisomeric CHF-phosphonates in an active site of interest) from a single primary triflate. The properties of the title compounds in enzyme active sites are discussed, as are possible interactions of these fluorine-containing bioisosteres with active site residues.

Use of a Robust Dehydrogenase from an Archael Hyperthermophile in Asymmetric Catalysis−Dynamic Reductive Kinetic Resolution Entry into (S)-Profens

Described is an efficient heterologous expression system for Sulfolobus solfataricus ADH-10 (Alcohol Dehydrogenase isozyme 10) and its use in the dynamic reductive kinetic resolution (DYRKR) of 2-arylpropanal (Profen-type) substrates. Importantly, among the 12 aldehydes tested, a general preference for the (S)-antipode was observed, with high ee’s for substrates corresponding to the NSAIDs (nonsteroidal anti-inflammatory drugs) naproxen, ibuprofen, flurbiprofen, ketoprofen, and fenoprofen. To our knowledge, this is the first application of a dehydrogenase from this Sulfolobus hyperthermophile to asymmetric synthesis and the first example of a DYRKR with such an enzyme. The requisite aldehydes are generated by Buchwald−Hartwig-type Pd(0)-mediated α-arylation of tert-butyl propionate. This is followed by reduction to the aldehyde in one [lithium diisobutyl tert-butoxyaluminum hydride (LDBBA)] or two steps [LAH/Dess−Martin periodinane]. Treatment of the profenal substrates with SsADH in 5% EtOH/phosphate buffer, pH 9, with catalytic NADH at 80 °C leads to efficient DYRKR, with ee’s exceeding 90% for 9 aryl side chains, including those of the aforementioned NSAIDs. An in silico model, consistent with the observed broad side chain tolerance, is presented. Importantly, the SsADH-10 enzyme could be conveniently recycled by exploiting the differential solubility of the organic substrate/product at 80 °C and at rt. Pleasingly, SsADH-10 could be taken through several “thermal cycles,” without erosion of ee, suggesting this as a generalizable approach to enzyme recycling for hyperthermophilic enzymes. Moreover, the robustness of this hyperthermophilic DH, in terms of both catalytic activity and stereochemical fidelity, speaks for greater examination of such archaeal enzymes in asymmetric synthesis.

Enantioselective, ketoreductase-based entry into pharmaceutical building blocks: ethanol as tunable nicotinamide reductant.

The use of NADH- and NADPH-dependent ketoreductases to access enantioenriched pharmaceutical building blocks is reported. Seven structurally diverse synthons are obtained, including those for atomoxetine (KRED 132), talampanel (RS1-ADH and CPADH), Dolastatin (KRED 132), and fluoxetine (KRED 108/132). Ethanol may be used as stoichiometric reductant, regenerating both nicotinamide cofactors, particularly under four-electron redox conditions. Its favorable thermodynamic and economic profile, coupled with its advantageous dual cosolvent role, suggests a new application for biomass-derived ethanol.

Synthesis of the (α,α-difluoroalkyl)phosphonate analogue of phosphoserine

Abstract The synthesis of the (α,α-difluoroalkyl)phosphonate analogue of L-phosphoserine, 5 , in a form appropriate for solid phase peptide synthesis, is reported. Two independent routes are described, starting from L-serine or (R)-isopropylideneglycerol. In each case, PCF 2 C bond formation is achieved by triflate displacement with diethyl lithiodifluoromethylphosphonate.

ENANTIOMERICALLY ENRICHED ALPHA -METHYL AMINO ACIDS. USE OF AN ACYCLIC, CHIRAL ALANINE-DERIVED DIANION WITH A HIGH DIASTEREOFACIAL BIAS

Hindered esters derived from N-benzoylalanine and the following chiral alcohols have been synthesized: (1) (-)-isopinocampheol; (2) (-)-trans-2-phenylcyclohexanol and (3) (-)-8-phenylmenthol. Sequential treatment of these esters with LDA (1.2 equiv.) and n-butyllithium (2.4 equiv.) at -78°C in THF generates the corresponding chiral dianions. Alkylation of each of these with benzyl bromide reveals that only the (-)-8-phenylmenthyl auxiliary confers a high diastereofacial bias upon its derivative dianion. In fact, that dianion (6) consistently displays diastereomeric ratios in the range of 89:11 to 94:6 for alkylations with a spectrum of nine alkyl halides. If one recrystallization step is included, a single diastereomeric product may be obtained, as is demonstrated for the benzylation of 6. Of particular note, the alkylation with 3,4-bis(tert-butyldimethylsilyloxy)benzyl bromide (18) (94:6 diast. ratio, 72% yield) constitutes a formal synthesis of the clinically important antihypertensive (S)-α-methyl-DOPA (Aldomet), in enantiomerically enriched from. In all cases studied, yields are markedly improved, yet diastereoselectivities unchanged, by the addition of 10% HMPA to the reaction milieu. The (-)-8-phenylmenthol chiral auxiliary is conveniently recovered via ester cleavage with KO2/18-crown-6, following alkylation. Complete deprotection affords enantiomerically enriched (S)-α-methyl amino acids, in all cases examined, indicating that dianion 6 displays a substantial bias in favor of si face alkylation. This sense of diastereoselection is consistent with a chain-extended, internal chelate model for the reactive conformation of the dianion.

Stereocontrolled synthesis of quaternary β,γ-unsaturated amino acids: chain extension of d- and l-α-(2-tributylstannyl)vinyl amino acids

A pair of diastereomeric (4S,5S)- and (4S,5R)-4-methoxycarbonyl-5-phenylselenomethyl-2-phenyl oxazolines, derived from L-vinylglycine, serve as precursors to protected, quaternary, L- and D-α-(2-tributylstannyl)vinyl amino acids, respectively, in three steps {(i) alkylative side chain installation, (ii) eliminative ring-opening and (iii) vinyl selenide to vinyl stannane interconversion}. The title compounds may be protodestannylated to the corresponding free, quaternary L- and D-vinyl amino acids. Alternatively, the 2-stannylvinyl α-branch (or the derivative 2-iodovinyl branch) may be exploited to access novel quaternary, L- and D-β,γ-unsaturated amino acids via a range of transition metal-mediated cross coupling reactions.

Organoselenium-based entry into versatile, α-(2-tributylstannyl)vinyl amino acids in scalemic form: A new route to vinyl stannanes [14]

Described herein is a synthetically malleable class of quaternary,R-(2-trialkylstannyl)vinyl amino acid (AA) building blocks with potential applications inde noVo peptide design and engineering. The stereocontrolled route to these AAs highlights the versatility of the phenylseleno group, acting to (i) mask a double bond, (ii) direct a low-temperature alkylation reaction, (iii) facilitate an alkene unmasking step, and (iv) mediate the introduction of a stannylvinyl group through a new substitution reaction that is expected to prove useful in other synthetic contexts. In recent years, there has been heightened interest in R-branched AAs, in general. As the free monomers, quaternary AAs bearing â,γ-unsaturation are potential suicide inactivators for AAprocessing enzymes. 1 When incorporated into peptides, quaternary AAs can be used to promote R-helical,2 310-helical, or â-turn4 secondary structures. They may also be site-specifically engineered into proteins. 5 They are useful building blocks for natural products6 or combinatorial libraries, 7 and generally enhance the proteolytic stability of their derivative peptides. 8 For all such applications, scalemicR-branched AAs are desirable. 9-11 The stereodivergent route detailed below allows one to access the Dor L-enantiomer at will, and adds a dimension of synthetic flexibility inherent in the stannylvinyl R-branch.

In situ enzymatic screening (ISES): a tool for catalyst discovery and reaction development.

Themove from the more deliberate, traditional approach to catalyst discovery to combinatorial approaches, has spurred great interest in the development of parallel-screening methods. As Crabtree and Loch recently put it, ideally one seeks TMan appropriate chemical sensor in a rapid parallel assay to detect rate and perhaps selectivity∫.[1] Herein, we describe the use of enzymes to report rapidly on reaction rate, for a set of parallel reactions. Seto and Abato recently described the use of enzymes to report on (enantio)selectivity.[2] From a broader perspective, these developments may be regarded as adding to the versatility of enzymes as tools for the organic chemist, an area that has seen remarkable expansion from asymmetric processing of unnatural substrates[3±5] and protecting-group cleavage under mild conditions,[6] to the creation of artificial enzymatic pathways.[7, 8] There is currently great interest in TMcombinatorial catalysis∫,[9] especially in transition-metal (TM) catalyzed reactions, for which reaction discovery and optimization often involve varying 1) the metal, 2) the ligand (type, structure, and stoichiometry), and 3) the substrate structure. By choosing such a model reaction, we sought both to establish proof of principle and to assess the ability of the screen to evaluate such variables one at a time. In our approach, the organic reaction under study is coupled, in situ, with an enzymatic reaction that permits continuous UV spectroscopic monitoring of the reaction. We term this approach ×in situ enzymatic screening× (ISES). This method is complementary to the previously communicated screening techniques,[10±19] in that it provides 1) evidence of product formation (not directly available using the elegant IR-thermography method of the Morken and Reetz groups)[11] and 2) relative rate profiles (not easily available with time-point detection systems employing gas[12] or liquid chromatography[10, 13] or mass spectrometry),[14] 3) without the need to alter the substrate, by installing a chromophore,[15] a fluorophore,[16] , or an azo-dye precursor.[17] Consonant with our interest in developing synthetic methodology toward densely functionalized , -unsaturated amino acids[20] as inhibitors of PLP-dependent (PLP pyridoxal phosphate) enzymes,[21] we chose a TM-mediated allylicamination reaction as our model reaction. We were influenced, in this regard, by an important precedent from Trost et al. ;[22a] scalemic vinylglycinol had been synthesized through Pd0-mediated allylic amination.[22] We set out to use ISES to identify other TMs, including less expensive ones, which were capable of catalyzing the intramolecular allylic-amination reaction illustrated in Scheme 1.[23] Success here would, in principle, validate the use of ISES in screening other variants of the allylic-displacement reaction.[24]

Facile installation of the phosphonate and (α,α-difluoromethyl)phosphonate functionalities equipped with benzyl protection

Abstract Under appropriate conditions, dibenzyl (lithiomethyl)phosphonate and dibenzyl (lithiodifluoromethyl)phosphonate displace primary triflates to provide convenient access to the corresponding phosphonates, carrying benzyl ester protecting groups. This approach is of particular advantage for phosphonate ester deprotection, which may be achieved by simple hydrogenolysis.

α-Vinyllysine and α-vinylarginine are time-dependent inhibitors of their cognate decarboxylases

(±)-α-Vinyllysine and (±)-α-vinylarginine display time-dependent inhibition of L-lysine decarboxylase from B. cadaveris, and L-arginine decarboxylase from E. coli, respectively. A complete Kitz-Wilson analysis has been performed using a modification of the Palcic continuous UV assay for decarboxylase activity.