Autocatalysis and Organocatalysis with Synthetic Structures

Autocatalysis and organocatalysis with synthetic structures

Seiji Kamioka, Dariush Ajami, and Julius Rebek Jr.1

The Skaggs Institute for Chemical Biology and Department of Chemistry, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037

Contributed by Julius Rebek, November 5, 2009 (sent for review September 8, 2009)

The discovery of ribozymes led to the proposal of an RNA world, proline itself requires extensive synthetic sequences, but creation where a single type of molecule was supposedly capable of self- of an imidazolidinone in the space between the recognition replication and chemical catalysis. We show here that both auto- elements as in 2a could be contemplated in a single step—the catalysis and organocatalysis can be engineered into a synthetic same step that assembles the autocatalyst. This functional group, structure. The compound is shown to selectively accelerate its devised by MacMillan and coworkers (16, 17) is a versatile own formation and catalyze either hydrogenation or nucleophilic organocatalyst that operates through reversibly formed covalent addition to α,β-unsaturated aldehydes. The observed reactivity intermediates (18). indicates that the components of a purported pre-RNA world In the experiment, 2a could be obtained from condensation of conceivably include smaller organic molecules. aldehyde 3 with thymine-derived amino acid 4a in hot benzene (Fig. 1 and SI Appendix). The desired cis 2a was accompanied imidazolidinone catalysis ∣ self-replication ∣ template effects by the trans isomer 2c. The N-methyl thymine derivatives 2b and 2d were also prepared and used as controls because their “ ” tudies of prebiotic chemistry raise questions concerning which inability to base pair abrogates molecular recognition during Sfunctions came nearer the origin of life—genetics or metabo- their reactions with the diaminotriazine subunit. lism (1). The discovery of the catalytic activity of ribozymes pro- The earliest evidence that recognition-based reactions were vided an answer in which both functions could be accounted for in involved in the final steps of the synthesis came from the yields an “RNA world” (2). Although no one doubted that RNA could of reactions involving closely related structures. Specifically, the 2a 2c carry genetic information, some 15 years lapsed before an RNA cis (38%) was obtained in higher yield than trans (18%), 2b 2d molecule was shown to self-replicate (3), and this type of catalysis whereas the N-methyl (19%) and (15%) were produced 2a has since been developed to an extraordinary level of efficiency in comparable amounts. The preference for was diminished (4). Short, self-complementary DNA strands were shown to self- in a polar solvent such as MeOH, which competes for hydrogen 4a 4b replicate in 1986 (5), and autocatalysis based on molecular recog- bonds, and was eliminated entirely in the reaction of and 5 nition was found in simplified organic molecules (6) and even with the ester , which lacks a hydrogen-bonding site complemen- 4a CHEMISTRY peptides (7). At the present time, many types of self-replicating tary to the imide function of (SI Appendix). structures are known (8). These systems function like their nu- A consistent and quantitative measure of autocatalysis was obtained through kinetic studies. The condensation reaction of cleic acid counterparts in that the self-complementary structures 3 4a operate as templates for their own formation. However, unlike with was analyzed by HPLC and the results are shown in ribozymes, the synthetic self-replicating systems are not known Fig. 2 Left. The initial rate for the coupling reaction (blue line) is shown with that of 0.25 equiv (red line) and 0.50 equiv (green to catalyze other chemical reactions. The wholly synthetic struc- 2a tures act as either organocatalysts or autocatalysts, but published line) cis added. The yields were corrected for the addition of the catalysts and are determined with respect to an internal stan- evidence for both activities in a single molecule is limited (9). This 2a research was undertaken to find molecules that could perform in dard. Clearly, accelerates its own formation. That this process both capacities, and we report here an imidazolidinone-based is grounded in self-recognition was shown by parallel studies trans 2c 2a Right compound and its properties as a catalyst and autocatalyst. monitoring as a function of added (Fig. 2 ). Here, no acceleration of the rate occurred; cis 2a is, accordingly, a self- Results and Discussion ish (8) replicator because it catalyzes its own formation but not To realize an autocatalytic/organocatalytic molecule, we made that of its structurally related diastereomer trans 2c. By compar- modifications in the structure of compound 1, a molecule ison, trans 2c also accelerates its own formation and, unexpect- 2a 2c that bears self-complementary hydrogen-bonding subunits—a edly, the formation of the cis as well (S.I.). Thus, trans is diaminotriazine and cyclic imide—that allow for dimerization a cooperative (19) replicator. The trans arrangement of alkyl in noncompeting organic media (Fig. 1). The arrangement of groups on an imidazolidinone is not expected to be advantageous 2c these subunits in 1 was earlier shown to allow its action as an for organocatalysis so trans was not tested in those capacities. 2a 2c autocatalyst (10): It acts as a template for its own synthesis, gath- Both cis and trans are stable under the reaction conditions; ering two subunits on its surface to facilitate the formation of the they neither interconvert nor racemize. amide bond. The insertion of a functional group known to cata- Taken together, these results show that autocatalysis based on — — lyze organic transformations into 1 was a prospect to be explored. recognition replication takes place during the formation of 2a We initially inserted a thiourea into the framework of 1 (11). the cis , and the likely process is illustrated in Fig. 3. The two 2a Although the thiourea function is a widely used organocatalyst complementary hydrogen-bonding sites of (diaminotriazine (12), its applications were limited in the present context: The and thymine) can assemble the starting materials into the same hydrogen bonding that accounts for the thiourea’s activity requires the use of solvents that also lead to tight dimerization Author contributions: S.K., D.A., and J.R. designed research; S.K. and D.A. performed of the template. This effect minimizes the fraction of the mono- research; S.K., D.A., and J.R. analyzed data; S.K., D.A., and J.R. wrote the paper. meric compound that operates as the organocatalyst. To avoid The authors declare no conflict of interest. this limitation we sought an organocatalytic function that oper- 1To whom correspondence should be addressed. Email: [email protected]. ates through reversible formation of covalent bonds, such as This article contains supporting information online at www.pnas.org/cgi/content/full/ the venerable proline-based catalysts (13–15). Insertion of 0912769107/DCSupplemental.

www.pnas.org/cgi/doi/10.1073/pnas.0912769107 PNAS ∣ January 12, 2010 ∣ vol. 107 ∣ no. 2 ∣ 541–544 Downloaded by guest on September 27, 2021 Fig. 1. Line drawings and synthesis of autocatalytic molecules. The previously reported self-replicator 1, was modified to contain an embedded imidazolidinone in 2a. The condensation of aldehyde 3 with thymine-derived amino acid 4a results in both the cis 2a and the trans 2c. The N-methyl derivatives (2b, 2d, and 4b) and aldehyde 5 lack the corresponding hydrogen bond motifs (recognition elements) and were used as controls.

termolecular complex 6 to template the reaction. This complex The imidazolidinone motif is also known to catalyze the cou- places the two reacting sites, aldehyde and amine, in close proxi- pling of indoles and α,β-unsaturated aldehydes in Friedel–Crafts mity and accelerates the rate of their condensation reaction. alkylations (21) (Fig. 5). Compound 2a was also effective in this The imidazolidinone site in the self-replicating molecule 2a is context: The addition of N-methylindole 10 to 7 was catalyzed by capable of activating α,β-unsaturated aldehydes via the reversible 2a and control experiments with 3 and 4a clearly illustrated that formation of iminium ions (20). This catalytic functional group the catalytic activity of 2a is not housed in its diaminopyridine or forms during the syntheses of the replicator. The organocatalytic imide functional groups. activity of 2a was demonstrated in a hydride transfer reaction In the present study, neither the catalyst nor the reaction con- (Fig. 4) to (E)-cinnamaldehyde 7. The substrate was reduced ditions are conventionally prebiotic, and the replication does not in high yield (91%) using 2a, the Hantzsch ester 8 as the hydride involve an “informational” oligomer though such synthetic struc- source and catalytic amount of an acid (DCl). The course of tures do exist and are capable of information transfer and pairing the reaction was followed with 1H NMR spectroscopy in which with RNA (22, 23). There are also synthetic cyclic phosphates the downfield doublet of cinnamaldehyde changed to a trip- embedded in chiral microenvironments that feature the same let of dihydrocinnamaldehyde 9; the corresponding signals of functional group as the phosphodiesters of RNA and DNA. imidazolidinone catalyst 2a did not change. No reduction was Although the cyclic phosphates are widely used organocatalysts observed in the absence of 2a. Neither the thymine-derived (24), the minimalist dinucleotide monophosphates counterparts amino amide 4a nor the aldehyde 3 acted as a reduction catalyst have, to our knowledge, yet to show activity in this regard. The under these conditions. departure for the cases at hand is the postulate that autocatalytic

Fig. 2. Kinetics of autocatalytic reactions. (Left) Appearance of cis compound 2a as a function of time: blue line, no additive; red line, 0.25 equiv product added; green line, 0.50 equiv product added. Initial concentrations were aldehyde 3 (8.0 mM) and amine 4a (9.6 mM) in benzene. (Right) Appearance of trans compound 2c as a function of time: blue line, no additive; red line, 0.25 equiv 2a added; green line, 0.50 equiv 2a added. Initial concentrations were aldehyde 3 (8.0 mM) and amine 4a (9.6 mM) in benzene.

542 ∣ www.pnas.org/cgi/doi/10.1073/pnas.0912769107 Kamioka et al. Downloaded by guest on September 27, 2021 Fig. 3. Autocatalytic reaction through a proposed termolecular complex. The recognition sites of 2a act through hydrogen bonding and stacking to provide a template for assembly 6. The reactive groups (circled) can be brought into proximity and the condensation reaction is accelerated. A modeled structure of the dimeric 2a.2a is also shown.

reproduction of catalytic capabilities (25) can be a useful and 5 (2 mM) in benzene (2.0 mL) at room temperature. After 2 d of stirring expanded definition of the chemistry at life’s origins. Compound at 70 °C, the reaction mixture was concentrated in vacuo. The residue was 2a acting as both autocatalyst and organocatalyst provides a purified by column chromatography on silica gel, eluting with 30% acetone working example. Such organocatalysts could even be envisioned in chloroform to afford the cis and trans imidazolidinones. The coupling to assist in building up the molecular components from which reaction of 3 and 4a gave 53% of cis 2a and 24% of trans 2c. The character- they are composed, for example, the synthesis of the pyrimidine ization data for 2a and 2c are as follows. Cis 2a: 1H NMR (600 MHz, DMSO-d6) δ 11.26 (s, 1H), 7.75 (s, 1H), 7.67 heterocyclic subunits featured here. Such compounds provide a 7 05 − 6 95 starting point for a “bottom up” approach to creating molecular (s, 1H), 7.39 (s, 1H), 7.38 (s, 1H), 7.25 (s, 1H), . . (m, 3H), 6.78 (d, J ¼ 6.9 Hz, 2H), 6.20 (s, 1H), 4.37 − 4.30 (m, 2H), 4.14 (d, J ¼ 14.9 Hz, complexity and establishing catalytic reaction cycles (26). There 1H), 4.09 − 4.00 (s, 1H), 3.85 − 3.80 (m, 2H), 3.75 (s, 3H), 1.71 (s, 3H), 1.60 is also the possibility for mutations (27) in these synthetic (s, 3H), 1.59 (s, 3H), 1.32 (s, 9H), 1.19 (s, 9H); 13C NMR (151 MHz, DMSO-d6) replicators. The ultimate aim is the generation of accessible δ 172.8, 171.8, 168.0, 165.1, 152.1, 147.4, 146.6, 146.0, 145.4, 142.8, 137.8, synthetic systems that are potential precursors to the RNA world. 131.2, 129.9, 128.6, 127.6, 127.2, 127.1, 126.4, 125.8, 124.8, 124.3, 123.6, 108.6, 67.8, 57.7, 49.9, 44.5, 35.3, 35.1, 35.0, 33.8, 32.2, 32.1, 31.2, 13.0; Materials and Methods high-resolution mass spectrometry (HRMS) [matrix-assisted laser desorption/ General Experimental. All reagents were obtained from commercial suppliers ionization–Fourier transform mass spectrometry (MALDI-FTMS): MHþ] calcu- and used without further purification. NMR spectra were recorded on a þ lated for C42H50N9O 744.3998, found 744.3975. Bruker DRX-600 spectrometer at 300 K. HPLC for analysis was performed 4 Trans 2c: 1H NMR (600 MHz, DMSO-d6) δ 11.28 (s, 1H), 7.61 − 7.55 (m, 2H), on a Hewlett–Packard HP-1100 series system with following gradients: 7.38 (d, J ¼ 2.0 Hz, 1H), 7.25 (s, 1H), 7.08 (d, J ¼ 1.9 Hz, 1H), 7.06 − 6.96 5–40% over 7 min, 40–100% over 25 min, and then 100% over 5 min (m, 3H), 6.82 (d, J ¼ 6.9 Hz, 2H), 6.78 (s, 4H), 6.18 (d, J ¼ 6.5 Hz, 1H), [CH3CN in H2O (0.1% TFA)], with the flow rate of 0.5 mL∕ min on a GL 4 19 − 4 12 4 12 − 4 08 J ¼ 16 0 CHEMISTRY Sciences, Inc. Inertsil-ODS-1HO, 4.6 × 150 mm column. Peak areas were . . (m, 2H), . . (m, 1H), 4.06 (s, . Hz, 1H), 3.69 (dd, J ¼ 7 1 J ¼ 9 8 integrated at 254 nm. The details of the synthetic procedures and character- . , 13.5 Hz, 1H), 3.44 (dd, . , 13.5 Hz, 1H), 1.71 (s, 3H), 1.59 13 izations of the aldehydes 3 and 5, as well as thymine-derived amino acids 4a (s, 6H), 1.33 (s, 9H), 1.17 (s, 9H); C NMR (151 MHz, DMSO-d6) δ 172.3, 171.8, and 4b are described in SI Appendix. 167.2, 164.4, 151.0, 146.4, 145.9, 145.3, 144.6, 142.5, 136.8, 130.3, 129.6, 127.8, 127.1, 126.6, 126.0, 124.8, 124.4, 124.0, 123.0, 121.9, 107.7, 57.0, 49.4, General Procedure for the Synthesis of Imidazolidinones 2. Thymine-derived 44.0, 34.5, 34.3, 34.2, 31.5, 31.4, 31.3, 31.3, 31.1, 12.0; HRMS (MALDI-FTMS: þ þ amino acid 4a or 4b (3 mM) was treated with a solution of aldehyde 3 or MH ) calculated for C42H50N9O4 744.3998, found 744.3982.

Fig. 4. The organocatalyzed reduction of cinnamaldehyde: red line, 0.2 equiv 2a; green line, 0.2 eqiv 3; blue line, 0.2 equiv 4a; black line, no additive.

Kamioka et al. PNAS ∣ January 12, 2010 ∣ vol. 107 ∣ no. 2 ∣ 543 Downloaded by guest on September 27, 2021 data for cis 2b, 12a, and 12b, as well as trans 2d, 12c, and 12d are described in SI Appendix.

General Procedure for Kinetic Studies of Autocatalyzed Coupling Reactions of Aldehyde 3 with Amino Acid 4a. A solution of aldehyde 3 (8.0 mM), thymine- derived amino acid 4a (9.6 mM), and an internal standard such as biphenyl (8.0 mM) in benzene was treated with (no additive, 0.25 equiv of 2a,or 0.50 equiv of 2a) at room temperature. While stirring at 70 °C, the mixture was periodically analyzed by HPLC through comparison of the peak areas of the internal standard with products (254 nm).

General Procedure for Competition Studies for Coupling Reactions of Aldehyde 3 or 5 with Thymine-Derived Amino Acids 4a and 4b. Thymine-derived amino acids 4a and 4b (8.0 mM each) were mixed with a solution of aldehyde 3 or 5 (8.0 mM) in benzene or methanol at room temperature. After 2 d of stirring at 70 °C, the mixture was analyzed by HPLC using the peak areas (254 nm).

General Procedure for Organocatalytic Hydride Reduction. The trans- cinnamaldehyde 7 (5.0 mM) was added to a solution of imidazolidinone 2a (1.0 mM), Hantzsch ester 8 (50.0 mM), and a catalytic amount of conc. DCl in CD2Cl2 at 0 °C. The reaction was allowed to come to room temperature, and the mixture was periodically analyzed by 1H NMR. Fig. 5. The organocatalyzed Friedel-Crafts alkylation: red line, 0.2 equiv 2a; green line, 0.5 eqiv 3; blue line, 0.2 equiv 4a; black line, no additive. General Procedure for Organocatalytic Friedel–Crafts Alkylations. The trans- cinnamaldehyde 7 (5.0 mM) and camphorsulfonic acid (50.0 mM) were added to a solution of imidazolidinone 2a (2.5 mM) and N-methylindole (50.0 mM) The stereochemistry of above compounds was determined from a 2D in CDCl3 at 0 °C. While stirring at room temperature, the mixture was 1 rotating frame nuclear Overhauser effect spectroscopy spectrum. An NOE analyzed by H NMR analysis. cross-peak was observed for the benzylic proton of the xanthene and the α-proton of the amino acid in cis 2a, whereas the trans 2c showed NOE ACKNOWLEDGMENTS. We are grateful to The Skaggs Institute for support cross-peaks for the benzylic proton of the xanthene with the β-proton of and Prof. A. Eschenmoser for encouragement and advice. S.K. is a Skaggs the amino acid and methyl protons at thymine. The yields and spectroscopy Postdoctoral Fellow.

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