Tetrahedron Letters. Vo1.32, No.41, pp 5725-5728,199l Printed in Great Britain

oo4o-4039/91 $3.00 + .oo Pergamon Press plc

The Synthesis of Dihenzofuran Based Diacids and Amino Acids Designed to Nucleate Parallel and Antiparallel P-Sheet Formation. P

Humberto Diaz and Jeffery W. Kelly *

Department of Chemistry, Texas A&M University College Station, Texas 77843-3255

Abstracg 4-Wtert -butyloxycarbonyl-2-aminoethyl-)-6-dibenzofuranpropionic acid (1) and 4,6-

dibenzofumndipmpionic acid (2> designed to nucleate antiparallel and parallel g-sheet formation,

respectively, have been synthesized in multi-gram quantities.

Our understanding of parallel and antiparallel g-sheet secondary struchue in peptides and proteins lags

significantly behind our comprehension of the a-helical sttucture because the development of p-sheet

model systems have proven elusive. The inability to predict the locations of the required reverse turns or

loops ( the turn problem ) and the tendency of g-sheets to aggregate, once formed, are impeding progress

towards a well characterized g-sheet model system. 1 Nucleation of sheet formation may involve the

formation of two intramolecular hydrogen bonds between residues i and i+3, hence, the torsion angles for

the intervening residues must be restricted and compatible with this hydrogen bonding geometry (Fig la).

Nucleation is most likely the slowest and energetically most unfavorable step involved in g-sheet folding.

We have begun to address the turn problem by employing an unnatural dibenzofuran-based reverse turn

having restricted conformational entropy and the appropriate geometry to nucleate hydrogen bond

formation between strands at the optimal 4.gSA distance. We have, therefore, synthesized a simple

conformationally restricted amino acid 1 and a diacid 2 in order to examine their potential to nucleate the

formation of antiparallel and parallel g-sheet structures, respectively.

t:R= COOH (a)

1 J (b)

FIGURE 1 (a) Reverse turn found in proteins. (b) Dibenzofuran p-turn mimic, designed to replace

residues i+l and i+2 in a normal Type I g-turn, incorporated into an antiparallel g-sheet structure.

8 Dedicated to the late Professor E.T. Kaiser

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We aspire to produce a spacer which will nucleate the folding of a linear sequence of ca

twelve amino acids into a g-sheet. The concept of using a conformationally restricted spacer with the

appropriate dimensions and geometry to increase the stability of a given conformation in peptides has

existed for over a decade.2 Nevertheless, no systematic studies concerning the stabilities of the resulting j3-

sheets have been reported. The large quantities of sheet nucleator necessary for such an endeavor required

that we develop a compound that could be easily prepared on a large scale (>lOg). Dibenzofuran-based

diacids and amino acids seemed optimal because of their ideal geometry and because they could easily be

rendered water soluble (owing to their susceptibility towards electmphilic aromatic substitution reactions at

the 2 and 8 positions, e.g. sulfonation reactions). The dibenzofuran-based turns also facilitate investigating

the influence of conformational entropy on the turn’s nucleating ability. Derivatives of the turns reported

here having one or two fewer methylene groups are being synthesized for this purpose. We considered

the simple phenoxathiin S-dioxide reverse turn mimic developed by Feigel for sheet nucleation; however,

recent NMK studies question its effectiveness as a g-turn mimic.3a Many other turn mimetics were

considered and some of them may be tested eventually; however, the large quantities of material needed for

the initial studies made their use impractical. 3b A simple dibenzofuran-based “g-mm” would complement

the “p-turns” synthesized by Kahn, which serve to orient side chain functional groups analogous to the

orientation found in natural g-turns, ( Fig la ).4 Our turn nucleating approach also is complementary to

the approach taken by Kemp and coworkers who have synthesized g-sheet templates which nucleate ft-

sheet structum by functioning as an artificial g-strand.5

33% overall yield

The potential antiparallel g-sheet nucleator 1 and the parallel g-sheet nucleator 2 were synthesized

easily in multi-gram quantities. The synthesis of 1 and 2 employed dibenzofuran, which was metallated

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and subsequently silylated with trimethylsilyl chloride twice to afford 4,6-bis(trimethylsilyl)dibenzofiuan

(3_).6a The ipsodirecting ability of the TMS group in electrophilic aromatic reactions sufficed to effect the

synthesis of 4,6diiododibenzofuran 4 from J employing I-Cl. 6b37 Interestingly, Friedel-Crafts

electrophilic aromatic substitution reactions were not successful for functionalizing positions 4 and 6 using

3 as starting material, instead, 2 and 8 substitution was observed.6 The Heck reaction was employed to

append the acrylate side chains at the 4 and 6 positions on the dibenzofuran skeleton to afford the 4,6-bis

(E )-propenoic ethyl ester 9.8 Hydrolysis of s followed by hydrogenation yielded the potential parallel

sheet nucleator, 4,6dibenzofurandipropionic acid (2> in an overall yield of 72% from dibenzofuran9

The key synthetic transformation in our synthesis of 1 involves self catalysis to ester@ one of the two

identical acid functionalities. A solution of 2 in EtOH was heated at reflux in the presence of 3A molecular sieves to afford selectively the mono-ethyl ester, 6 ( < 9% diester by HPLC).Iu The mono-ester

6 was treated with diphenylphosphoryl azide in tertiary butyl alcohol to afford the tert -butyl carbamate 2

in 75% yield. II Selective cleavage of the ester via saponification yielded the Boc-protected amino acid salt

1. The overall yield of this seven step procedure was 33% from dibenzofuran (45% if the remaining

starting material in the preparation of 6 was recycled once).9 Preparations of 1 and 2 in log quantities

have been accomplished without any special procedures.

Amino acid 1 has been utilized in sequential solid phase peptide synthesis and in solution segment

condensations to make pentapeptides and nonapeptides containing 1. Diacid 2 has been coupled in

solution to protected peptides to form both penta- and nonapeptides. Biophysical studies of the

conformation of peptides containing residues 1 and 2 are now in progress. Details of the these studies

including detailed procedures for the synthesis of 1 and 2 and particulars concerning the synthesis of

peptides containing 1 and 2, will be reported in due course.

Acknowl~: Financial support from the Robert A. Welch Foundation, the National Institute of

Health Biomedical Research Support Grant Program, and the Searle Scholars Program / The Chicago

Community Trust (JWK) is gratefully acknowledged. We also thank the referees for the insightful review.

REFERENCES and NOTES

1. See Creighton, T.E. “Proteins-Structures and Molecular Principles” W.H. Freeman and Co., New York, 1984 , page 191.

2. 3.

M.E.;Hill,D.E.; Kahn, M.; Madison, V.S.; Cook, C. M. J. Am. Cherit. Sot. 1990,112, 323.

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_ ,, L

4. Kahn, M.; Bertenshaw, C. M. Tettrahedron Lett. 198 /

.B j ,2317-233@ Kahn, M.;

Chen, C. M. Tetrahedron L&t. 1987,

P

1623-1626 ?A n, M.; Devens, B. Tetrahedron L.&t. 1986,27,4841-44, hn, M.; Wilke, S.; Chen, B.; Fuji@ K. J. Am. Chem. Sot. 1988,110, 1638 1639.

5. Kemp, D.S.; Bowen, B.R. Tetrahedron Lett. 1988,29, 5 emp, D.S.; Bowen, B.R.; Muendel, C.C. J. Org. Chem. 1990,55,4650-4657

9

7-5080; Kemp, k .S. Trends in

Biotechnology 1990,6, 246-255. 6. a. Sargent, M. V.; &an&y, P.O. Adv. Heterocyclic Chem. 1984,35, l-80.

b. It was pointed out to us that Professor Cram’s group has synthesized 4 in one step from dibenzofuran in 65% yield. Benaksas Schwartz, E.J. Ph. D. Dissertation, University of California, Los Angeles, CA., 1989.

7. Chan, T.H.; Fleming, I. Synthesis 1979, 761-786.; Hillard, R.L.; Vollhardt, K.P.C. J. Am. Gem. Sot. 1977,99, 4058.

8. Heck, R. F.; Melpolder, J. B. J. Org. Chem. 1976.41, 265. 9. Compounds 1-z were characterized by lH NMR and l3C NMR spectroscopy, by

elemental analysis, and in the case of 2, by X-ray diffraction, limited data below.

3; mp 83-84 OC; lHNMR (Acetone-&) 6 0.48 (s, 9 H, Si -CH3), 7.37 (t, J =7.2 Hz, 1 H, C2 - H), 7.57 (dd, J =7.2 Hz, 1.4 Hz, 1 H, C3 - H), 8.11 (dd, J = 7.6 Hz, 1.4 Hz, 1 H, Cl - H); Anal. Calcd. for ClsHaOSi2: C, 69.17; H, 7.74. Found: C, 69.12; H, 7.89. 4; mp 160-162oC (lit. 160-161cC); lHNMR (Acetone-dg) 6 7.25 (t, J=7.7 Hz, lH, C2-H), 7.95 (dd, J=7.8 Hz,l.l Hz, lH, C3-H), 8.13 (dd, J=7.7 Hz, 1.1 Hz, lH, Cl-H); Anal. Calcd. for Cl$I&O: C, 34.32; H, 1.44. Found: C, 34.46; H, 1.41.

& mp 143-143.5 oC; lHNMR (Acetone-k) 8 1.37 (t, J =7.1 Hz, 3H, CH3), 4.30 (q, J =7.1 Hz, 2H, CH2), 7.01 (d, J = 16.2 Hz, lH, vinylic proton) 7.49 (t, J =7.6 Hz, lH, C2 - H), 7.84 (dm, J = 7.6 Hz, 2H, C3 - H), 8.01 (d, J = 16.3 Hz, lH, vinylic proton), 8.19 (dd, J = 7.7 Hz, 1.24 Hz, lH, Cl - H); Anal. Calcd. for C22H2005: C, 72.51; H, 5.53. Found: C, 72.50; H, 5.46.

2; mp 204-206oC *HNMR (DMSG-de) 6 12.20 (s, OH), 7.95 (dd, J=7.3 Hz, 1.6 Hz, lH, Cl-H), 7.37 (dd, J=7.4 Hz, 1.6 Hz, lH, C3-H), 7.29 (t, J=7.4 Hz, IH, C2-H), 3.19 (t, J=7.7 Hz, CH2) 2.75 (t, J=7.8 Hz, CH2); Anal. Calcd. for ClgHl605: C, 69.22; H, 5.16. Found: C, 68.97; H, 4.87.

6; mp 92-93 oC; lHNMR (Acetone&j) 6 10.65 (bs, OH), 7.92 (dd, J=7.4 Hz, 1.5 Hz, 2H, C1,9- H), 7.39 (m, 2H), 7.30 (m, 2H), 4.07 (q, J=7.1 Hz, 2H, 0-CH2), 3.31 (t, J=7.6 Hz, 4H, 2CH2 ), 2.85 (m, 4H, 2CH2), 1.16 (t, J=7.1 Hz, 3H, CH3). Anal. Calcd. for C2OH2OO5: C, 70.58; H, 5.92. Found: C, 70.57; H, 5.90.

2; ‘HNMR (Acetone-d6) 8 7.92 (dd, J = 7.4 Hz, 1.6 Hz, 2H, C1,9-H) 7.54 (m, 4H), 6.15 (bs, lH, N-H), 4.08 (q, J=7.12 Hz, 0-CH2) 3.55 (q, J = 6.2 Hz, 2H, CH2-NH) 3.31 (t, J = 7.9Hz, 2H, CH2) 3.19 (t, J = 7.3 Hz, 2H, CH2) 2.85 (t, J = 7.8 Hz, 2H, CH2) 1.34 (s. 9H, t-butyl) 1.17 (t, J = 7.1 Hz, 3H, CH3); Anal. Calcd. for C24H2gN05: C, 70.05; H, 7.10. Found: C, 70.25; H, 7.14.

10. A linear intramolecular hydrogen bond 2.66A in length (2.65 f 0.5A is ideal) is geometrically possible, as evidenced by minimizing the X-ray structure of 2 with the MM2 force field in vacuum ( Macromodel ). In MeOH the electrostatic interactions are much less important, hence, invoking intramolecular catalysis as the origin of the observed catalysis is not possible without further experimental evidence. For an lead reference on Maromodel see: Mohamadi, F.; Richards, N.G.J.; Guida, W.C.; Liskamp, R.; Lipton, M.;Caufield,C.; Chang, G.; Hendrickson, T.; Still, W.C. J. Comput. Chem. 1990,11, 440-467.

11. Haefliger, W.; Kloppner, E. Helv. Chim. Acta 1982,65, 1837.; Shioiri, T.; Ninomiya, S.; Yamada, S. J. Am. Chem. Sot. 1972,94, 6203.

(Received in USA 24 April 1991)