In Situ Enzymatic Screening (ISES): A Tool for Catalyst Discovery and Reaction Development

Abstract

Dedicated to Professor Samuel J. Danishefsky on the occasion of his 65th birthdayThe move 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 ™an 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 ™combinatorial cataly-sis∫, [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 b,g-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 Pd 0 -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] Scheme 1. ISES of catalysts for allylic displacement. PMP para-methoxyphenyl.

The In Situ Enzymatic Screening (ISES) Approach to Reaction Discovery and Catalyst Identification

“…For example, the product must diffuse into the aqueous reporting phase readily enough to obtain useful initial rate information. Adjusting the solvent composition may facilitate product diffusion and can also limit the rate of water diffusion into the organic layer (Berkowitz et al., ). Proper solvent choice also limits precipitation at the organic/aqueous interface, a significant issue when an upper organic layer is used.…”

“…The first iteration of ISES was developed to explore catalytic combinations for a transition metal‐mediated intramolecular amination reaction upon an allylic ethyl carbonate substrate (Berkowitz & Maiti, ; Berkowitz et al., ; Berkowitz, Bose, & Choi, ; Figure , ). Turnover of such substrates is expected to proceed via formation of a π‐allyl or σ‐allyl‐metal intermediate with release of the ethyl carbonate leaving group in each catalytic cycle.…”

“…( E ) Linear least squares fit rates of NADH formation for the catalytic combinations being sampled. Elements of this figure were adapted from Berkowitz et al., with permission from Wiley‐VCH Verlag. ADH, alcohol dehydrogenase; AlDH, aldehyde dehydrogenase; PMP, p ‐methoxyphenyl; TM, transition metal.…”

“…This is an example of the use of ISES for air‐sensitive chemistry, as can be seen in Figure B; the cuvettes have all been purged with Ar, and are maintained under an inert atmosphere via septa. Indeed, the combination of p ‐methoxyphenyl (PMP) protection on the nucleophilic nitrogen, Ni(cod) 2 , and bidentate phosphine ligands was found to be optimal based solely on iterative ISES screening (Berkowitz et al., ). This ultimately led to the first examples of asymmetric Ni(0)‐mediated allylic amination chemistry and to the first asymmetric base metal‐mediated synthesis of L ‐vinylglycine (Berkowitz & Maiti, ; Berkowitz et al., ).…”

Combinatorial Catalysis Employing a Visible Enzymatic Beacon in Real Time: Synthetically Versatile (Pseudo)Halometalation/Carbocyclizations

“…Our group has developed an in situ enzymatic screening (ISES) approach whereby an organometallic reaction under study is coupled to an enzymatic reporting reaction in real time. [17] This screening method led to the discovery of the first asymmetric allylic amination with nickel(0) [18] and to the identification of novel salen [salen = N,N'-bis(salicylidene)ethylenediamine)] ligands with promise for asymmetric synthesis. [19] Those approaches involved dehydrogenase enzymes [20] as sensors, thus utilizing the inherent nicotinamide cofactor to provide a UV signal.…”