Incorporation of Phosphole Moieties into the Side Chain of Tyrosine and Phenylalanine

Abstract

A tyrosine derivative with a phosphole moiety covalently attached to the phenolic hydroxy group was successfully prepared through P-O bond-forming reaction, via a nucleophilic substitution reaction at the phosphorus, by slow addition of N-CBz-protected tyrosine methyl ester to a chlorophosphole adduct in the presence of triethylamine, albeit with a low yield. Furthermore, a phenylalanine derivative with a phosphole-containing side chain was conveniently synthesized by Stille cross-coupling reaction of a stannylphosphole reagent with N-Boc-protected 4-iodophenylalanine methyl ester, using Pd(dba)₂ as catalyst, in the absence of any additional ligand. These molecules must be seen as valuable building blocks for the preparation of metalloproteins of interest for chemical, biological, and medical applications.

α-amino acids - phosphine-containing side chain - design - functional metalloproteins - phosphole chemistry - P-O bond-forming reaction - Stille reaction

References and Notes

1

Deceased March 17, 2010

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Z-Tyr-OH (97%) was purchased from Sigma-Aldrich Co.

22

Experimental Procedure for N -CBz-Protected l -Tyrosine Methyl Ester (1): The Schlenk technique was used. To a solution of Z-Tyr-OH (3.0 g, 9.51 × 10^-³ mol)
in DMF (23.8 mL → c = 0.4 M) at 0 ˚C were added subsequently KHCO₃ (1.05 g, 1.05 × 10^-² mol) and MeI (1.18 mL, 1.90 × 10^-² mol). The reaction mixture was stirred overnight at r.t. The reaction was quenched with a sat. aq solution of NaHCO₃. The mixture was extracted with EtOAc, dried over MgSO₄ and concentrated in vacuo. Purification of the residue by silica gel chromatography (EtOAc-hexane, 1:2; deposit in CH₂Cl₂) allowed isolation of the desired compound (3.0 g, 9.12 × 10^-³ mol, yield: 96%) as a white solid. The analytical data were consistent with the previously reported data for this compound (Cf. ref. 18).

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24

Experimental Procedure for 1-Chloro-2,3,4,5-tetraphenylphosphole (2): The synthetic procedure was adapted from ref 34a. The Schlenk technique was used. n-BuLi (8.6 mL, c = 1.6 M, 1.37 × 10^-² mol) was added to a solution of zirconocene dichloride (Cp₂ZrCl₂, 2.0 g, 6.84 × 10^-³ mol) in THF (27.3 mL → c = 0.25 M) at -78 ˚C. The resulting reaction mixture was stirred for 1 h at -78 ˚C. After treatment of (n-Bu)₂ZrCp₂ generated in THF with diphenylacetylene (2 equiv, 2.44 g, 1.37 × 10^-² mol), the temperature of the reaction mixture was allowed to rise to r.t. overnight. After evaporation of THF, CH₂Cl₂ was added and the reaction mixture was filtered. The solvents were evaporated and CH₂Cl₂ was added (22.8 mL → c = 0.3 M). Phosphorus trichloride (597 µL, 6.84 × 10^-³ mol) was subsequently added at 0 ˚C. The temperature of the reaction mixture was allowed to rise to r.t. After 30 min, the solvents were removed in vacuo. A solution of hexane-THF (1:1) was added and the reaction mixture was filtered. The solvents were evaporated to produce the desired compound quantitatively. ^³¹P{^¹H} NMR of the crude reaction mixture in CDCl₃ showed one major signal at δ = 76.4 ppm (s). No purification was attempted on this substrate.

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31

Boc-Phe(4-I)-OH [≥99.0% (TLC)] was purchased from Sigma-Aldrich Co.

32

Experimental Procedure for N -Boc-Protected l -4-Iodophenylalanine Methyl Ester (5): The Schlenk technique was used. To a solution of Boc-Phe(4-I)-OH (3.0 g, 7.67 × 10^-³ mol) in DMF (19.2 mL → c = 0.4 M) at 0 ˚C were added subsequently KHCO₃ (845 mg, 8.44 × 10^-³ mol) and MeI (0.955 mL, 1.53 × 10^-² mol). The reaction mixture was stirred overnight at r.t. The reaction was quenched with a sat. aq solution of NaHCO₃. The mixture was extracted with EtOAc, dried over MgSO₄ and concentrated in vacuo. Purification of the residue by silica gel chromatography (EtOAc-hexane, 1:3) allowed isolation of the desired compound (3.0 g, 7.41 × 10^-³ mol, yield: 96%) as a white solid. The analytical data were consistent with the previously reported data for this compound (Cf. ref. 29).

33a

Experimental Procedure for 3,4-Dimethyl-2-phenyl-1-tributylstannylphosphole (6): The synthetic procedure was adapted from ref. 27b. The Schlenk technique was used. To potassium 3,4-dimethyl-2-phenylphospholidediglyme adduct (prepared according to ref. 33b; 1.0 g, 2.77 × 10^-³ mol) in THF (4.62 mL → c = 0.6 M), at 0 ˚C, tributyltin chloride (752 µL, 2.77 × 10^-³ mol) was added. After stirring at r.t. for 5 min, the solvent was distilled off. Degassed hexane was added. The mixture was filtered into another Schlenk flask via a cannula. The solid residue was washed with degassed hexane (2 ×). The hexane was distilled off to give the desired product as a yellow oil (conversion: 100%). ^³¹P{^¹H} NMR of the crude reaction mixture in CDCl₃ showed one single signal at δ = -50.1 ppm (s). This product was dissolved in THF (13.8 mL) to produce a 0.2 M solution of 3,4-dimethyl-2-phenyl-1-tributylstannylphosphole.

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36

General Procedures: All reactions were routinely performed under an inert atmosphere of argon or nitrogen by using Schlenk and glovebox techniques and anhyd deoxygenated solvents. Anhyd THF and hexane were obtained by distillation from Na/benzophenone. Anhyd CH₂Cl₂ was distilled over P₂O₅. NMR spectra were recorded at r.t. on a Bruker AC-300 SY spectrometer operating at 300.0 MHz for ^¹H NMR, 75.5 MHz for ^¹³C NMR, and 121.5 MHz for ^³¹P NMR. Solvent peaks are used as internal reference relative to Me₄Si for ^¹H NMR and ^¹³C NMR chemical shifts (ppm). ^³¹P chemical shifts are relative to an 85% H₃PO₄ external reference. Coupling constants are given in Hz. The following abbreviations are used: s, singlet; d, doublet; t, triplet; m, multiplet. All reagents and chemicals were obtained commercially and used as received.
Experimental Procedure for ( S )-2-Benzyloxy-carbonylamino-3-[4-(2,3,4,5-tetraphenyl-1-thioxo-phosphol-1-yloxy)phenyl]propionic Acid Methyl Ester (3): The Schlenk technique was used. A solution of N-CBz-protected l-tyrosine methyl ester (1.125g, 3.42 × 10^-³ mol) and Et₃N (477 µL, 3.42 × 10^-³ mol) in THF (68.4 mL → c = 0.05 M) was added dropwise (over 4-5 h) to the starting 1-chloro-2,3,4,5-tetraphenylphosphole (2.89 g, 6.84 × 10^-³ mol) in THF (68.4 mL → c = 0.1 M) at -78 ˚C. The temperature of the reaction mixture was allowed to rise to r.t. for 12 h. The white solid was filtered off and washed with Et₂O. The filtrates and washings were combined and the solvent was removed under reduced pressure. ^³¹P{^¹H} NMR of the crude reaction mixture in CDCl₃ showed one major signal at δ = 120.0 ppm (s) and a small side product at δ =
-12.0 ppm (s). The crude product was dissolved in THF (22.8 mL → c = 0.15 M) and elemental sulfur (438 mg,
1.37 × 10^-² mol) was added. The reaction mixture was stirred for 48 h at r.t. An aliquot was taken from the crude reaction mixture for analysis (conversion: 100%). ^³¹P{^¹H} NMR of the aliquot (in THF) showed no more signal at δ = 120.0 ppm, one new signal at δ = 93.8 ppm (s), and a minor signal (less than 5% conversion) at δ = 78.3 ppm (s). The solvents were removed under reduced pressure. Purification of the residue by silica gel chromatography (hexane-EtOAc, 3:1; deposit CH₂Cl₂) allowed isolation of the desired compound (515 mg, 6.89 × 10^-4 mol, yield: 20%) as a yellow oil, which slowly solidified under vacuum to give a light yellow glass. ^¹H NMR (300.131 MHz, CDCl₃): δ = 2.86-3.04 (2 H, m), 3.58 (3 H, s), 4.47-4.56 [1 H, 2 ¥ t overlapping (2^nd order signal), ^³ J = 6.3 Hz], 4.99 (2 H, s), 5.06 (1 H, 2 × br s overlapping), 6.60-7.40 (29 H, m). ^¹³C NMR (75.468 MHz, CDCl₃): δ = 39.3 (s), 54.2 (s), 56.5 (s), 68.9 (s), 123.9 (d, J _P = 4.0 Hz), 129.9-130.1 (m), 130.4 (s), 131.4 (s), 131.8 (d, J _P = 1.6 Hz), 131.9 (d, J _P = 6.1 Hz), 133.1 (d, J _P = 95.4 Hz), 133.9 (d, J _P = 10.7 Hz), 134.6 (s), 136.3 (d, J _P = 18.9 Hz), 138.0 (s), 150.5 (d, J _P = 30.5 Hz), 151.2 (d, J _P = 11.8 Hz), 157.4 (s), 173.5 (s). ^³¹P{^¹H} NMR (121.497 MHz, CDCl₃): δ = 99.4 (s). HRMS (EI+): m/z [M⁺] calcd for C₄₆H₃₈O₅NSP: 747.2208; found: 747.2237.
Experimental Procedure for ( S )-2- tert -Butoxy-carbonylamino-3-[4-(3,4-dimethyl-2-phenyl-1-thioxophosphol-1-yl)phenyl]propionic Acid Methyl Ester (8): The Schlenk technique was used. N-Boc-protected l-4-iodophenylalanine methyl ester (849 mg, 2.10 × 10^-³ mol) was dissolved in a solution of 3,4-dimethyl-2-phenyl-1-tributylstannylphosphole (10.5 mL, 0.2 M, 2.10 × 10^-³ mol) in THF. Pd(dba)₂ (24 mg, 4.19 × 10^-5 mol, 2 mol%) was added and the reaction mixture was stirred for 12 h at 60 ˚C. After cooling off the reaction mixture to r.t., an aliquot was taken (conversion: 100%). ^³¹P{^¹H} NMR of the aliquot (in THF) showed one signal at δ = 5.2 ppm (s). Elemental sulfur (135 mg, 4.19 × 10^-³ mol) was added and the reaction mixture was stirred at r.t. for 24 h. An aliquot was then taken from the crude reaction mixture for analysis (conversion: 100%). ^³¹P{^¹H} NMR of the aliquot (in THF) showed no more signal at δ = 5.2 ppm, one new signal at δ = 49.8 ppm (s). The solvents were removed under reduced pressure. Purification of the residue by silica gel chroma-tography (hexane-EtOAc, 3:1) allowed isolation of the desired compound (814 mg, 1.64 × 10^-³ mol, yield: 78%) as a 1:1 mixture of its two diastereoisomers, as a white solid. ^¹H NMR (300.131 MHz, CDCl₃): δ = 1.33 (9 H, s), 2.02 (3 H, d, J _P = 2.3 Hz), 2.12-2.16 (3 H, m), 2.90-3.10 [2 H, 2 × dd (2^nd order signals), ^³ J = 6.1 Hz, ^² J = 13.8 Hz], 3.55-3.59 (3 H, 2 × br s overlapping, 1:1 ratio), 4.40-4.55 [1 H, 2 × t overlapping (2^nd order signal), ^³ J = 6.3 Hz], 4.90 (1 H, 2 × br s overlapping), 6.11 (1 H, d, J _P = 31.0 Hz), 7.08 (2 H, dd,
J _P = 2.3 Hz, ^³ J = 8.0 Hz), 7.12-7.24 (5 H, m), 7.64 (2 H, dd, ^³ J = 8.2 Hz, J _P = 13.7 Hz). ^¹³C NMR (75.468 MHz, CDCl₃): δ = 13.6 (d, J _P = 14.1 Hz), 17.2 (d, J _P = 17.2 Hz), 27.3 (s), 37.4 (m), 51.3 (s), 53.2 (m), 79.1 (s), 122.6 (d, J _P = 83.3 Hz), 125.4 (dm, J _P = 77.3 Hz), 126.9 (d, J _P = 1.4 Hz), 127.3 (s), 127.9 (d, J _P = 4.6 Hz), 128.7 (d, J _P = 12.9 Hz), 129.9 (d, J _P = 12.1 Hz), 131.8 (d, J _P = 11.8 Hz), 136.5 (dm, J _P = 80.5 Hz), 139.7 (d, J _P = 2.9 Hz), 145.3 (dm, J _P = 24.4 Hz), 153.7 (d, J _P = 16.9 Hz), 153.9 (m), 170.87, 170.92 (2 × s). ^³¹P{^¹H} NMR (121.497 MHz, CDCl₃): δ = 54.9 (s). HRMS (EI+): m/z [M+] calcd for C₂₇H₃₂O₄NSP: 497.1790; found: 497.1779.