Please cite this article in press as: Kojima et al., Cyclization Reaction-Based Turn-on Probe for Covalent Labeling of Target Proteins, Cell Chemical
Biology (2020), https://doi.org/10.1016/j.chembiol.2020.01.006
MHz, CDCl
3
) d [ppm]: 174.1, 163.1, 160.2, 154.2, 136.0, 107.6, 106.1, 104.6, 86.2, 83.5, 71.9, 63.2, 61.9, 56.6, 56.1, 55.5, 42.8, 42.2,
4
0.5, 40.3, 36.6, 35.9, 35.5 (2 carbons), 34.7, 31.2, 31.0, 30.7, 28.3, 27.3, 26.5, 25.7 (3 carbons), 24.3, 23.5, 20.9, 18.4, 18.3, 12.1, -4.3
+
Si [M + H] 709.44995, found 709.45047.
(
2 carbons). HRMS (ESI) caled. for C42
H
65
O
7
2
-(2-((3-(2-((tert-Butyldimethylsilyl)oxy)-4-methoxyphenyl)propioloyl)oxy)ethoxy)ethyl (R)-4-
((3R,5R,8R,9S,10S,13R,14S,17R)-3-Hydroxy-10,13-dimethylhexadecahydro-1H-cyclopenta[a]phenanthren-17-yl)
pentanoate (40)
To a solution of 33 (47.2 mg, 0.15 mmol) and 35 (63.5 mg, 0.14 mmol) in CH
2
Cl
DCC (31.8 mg, 0.15 mmol) at 0 C and the mixture was stirred at room temperature for 16.5 h. The crude solution was purified by flash
2
(1.4 mL) was added DMAP (1.7 mg, 0.014 mmol) and
ꢀ
1
column chromatography (silica gel: 10 g, 45% AcOEt/hexane) to give 40 (45.5 mg, 43%). H NMR (300 MHz, CDCl
d, J = 8.6 Hz), 6.52 (1H, dd, J = 8.6, 2.4 Hz), 6.37 (1H, d, J = 2.4 Hz), 4.37-4.33 (2H, m), 4.25-4.22 (2H, m), 3.91-3.48 (8H, overlapped),
3
) d [ppm]: 7.43 (1H,
13
2
.45-2.20 (2H, m), 1.96-1.09 (27H, overlapped), 1.04 (9H, s), 0.91-0.89 (6H, overlapped), 0.63 (3H, s), 0.26 (6H, s). C NMR (75 MHz,
CDCl
3
) d [ppm]: 174.4, 163.0, 160.2, 154.4, 135.9, 107.6, 106.2, 104.7, 85.9, 83.7, 72.0, 69.3, 68.9, 64.6, 63.4, 56.6, 56.1, 55.6, 42.8,
4
1
2.2, 40.5, 40.3, 36.6, 36.0, 35.5 (2 carbons), 34.7, 31.3, 31.0, 30.7, 28.3, 27.3, 26.5, 25.8 (3 carbons), 24.3, 23.5, 20.9, 18.38, 18.35,
+
8
2.2, -4.3 (2 carbons). HRMS (ESI) caled. for C44H68NaO Si [M + Na] 775.45811, found 775.46107.
2
-(2-(2-((3-(2-((tert-Butyldimethylsilyl)oxy)-4-methoxyphenyl)propioloyl)oxy)ethoxy)ethoxy)ethyl (R)-4-
((3R,5R,8R,9S,10S,13R,14S,17R)-3-Hydroxy-10,13-dimethylhexadecahydro-1H-cyclopenta[a]phenanthren-17-yl)
pentanoate (41)
To a solution of 33 (37.1 mg, 0.12 mmol) and 36 (55.0 mg, 0.11 mmol) in CH
2
Cl
DCC (25.0 mg, 0.12 mmol) at 0 C and the mixture was stirred at room temperature for 20.5 h. The crude solution was purified by flash
2
(1.1 mL) was added DMAP (1.3 mg, 0.011 mmol) and
ꢀ
1
column chromatography (silica gel: 10 g, 55% AcOEt/hexane) to give 41 (41.0 mg, 47%). H NMR (300 MHz, CDCl
d, J = 8.6 Hz), 6.52 (1H, dd, J = 8.6, 2.4 Hz), 6.37 (1H, d, J = 2.4 Hz), 4.37-4.34 (2H, m), 4.24-4.21 (2H, m), 3.91-3.45 (12H, overlapped),
3
) d [ppm]: 7.42 (1H,
13
2
.44-2.17 (2H, m), 1.97-1.10 (27H, overlapped), 1.04 (9H, s), 0.91-0.89 (6H, overlapped), 0.63 (3H, s), 0.26 (6H, s). C NMR (75 MHz,
CDCl
3
) d [ppm]: 174.4, 163.0, 160.1, 154.4, 135.9, 107.6, 106.2, 104.7, 85.8, 83.7, 71.9, 70.8, 70.7, 69.4, 69.0, 64.6, 63.5, 56.6, 56.1,
5
1
5.5, 42.8, 42.2, 40.5, 40.3, 36.6, 36.0, 35.5 (2 carbons), 34.7, 31.3, 31.0, 30.7, 28.3, 27.3, 26.5, 25.8 (3 carbons), 24.3, 23.5, 20.9,
+
73 9
8.4, 18.3, 12.1, -4.3 (2 carbons). HRMS (ESI) caled. for C46H O Si [M + H] 797.50238, found 797.49795.
1
5-(2-((tert-Butyldimethylsilyl)oxy)-4-methoxyphenyl)-13-oxo-3,6,9,12-tetraoxapentadec-14-yn-1-yl (R)-4-
((3R,5R,8R,9S,10S,13R,14S,17R)-3-Hydroxy-10,13-dimethylhexadecahydro-1H-cyclopenta[a]phenanthren-17-yl)
pentanoate (42)
To a solution of 33 (47.2 mg, 0.15 mmol) and 37 (80.0 mg, 0.14 mmol) in CH
2
Cl
2
(1.4 mL) was added DMAP (1.7 mg, 0.014 mmol) and
ꢀ
DCC (31.8 mg, 0.15 mmol) at 0 C and the mixture was stirred at room temperature for 19 h. The crude solution was purified by flash
1
column chromatography (silica gel: 10 g, 60% AcOEt/hexane) to give 42 (58.4 mg, 50%). H NMR (300 MHz, CDCl
d, J = 8.6 Hz), 6.52 (1H, dd, J = 8.6, 2.4 Hz), 6.37 (1H, d, J = 2.4 Hz), 4.36-4.33 (2H, m), 4.23-4.20 (2H, m), 3.91-3.49 (16H, overlapped),
3
) d [ppm]: 7.42 (1H,
13
2
1
4
1
3
.43-2.17 (2H, m), 1.04 (9H, s), 0.91-0.89 (6H, overlapped), 0.63 (3H, s), 0.26 (6H, s). C NMR (75 MHz, CDCl ) d [ppm]: 174.4, 163.0,
60.1, 154.4, 135.9, 107.6, 106.2, 104.7, 85.8, 83.7, 71.9, 70.79, 70.77, 70.74, 70.70, 69.3, 69.0, 64.7, 63.5, 56.6, 56.1, 55.5, 42.8,
2.2, 40.5, 40.3, 36.6, 36.0, 35.5 (2 carbons), 34.7, 31.3, 31.0, 30.7, 28.3, 27.3, 26.5, 25.8 (3 carbons), 24.3, 23.5, 20.9, 18.4, 18.3,
+
2.2, -4.3 (2 carbons). HRMS (ESI) caled. for C48
H
77
O
10Si [M + H] 841.52860, found 841.52889.
1
8-(2-((tert-Butyldimethylsilyl)oxy)-4-methoxyphenyl)-16-oxo-3,6,9,12,15-pentaoxaoctadec-17-yn-1-yl (R)-4-
((3R,5R,8R,9S,10S,13R,14S,17R)-3-Hydroxy-10,13-dimethylhexadecahydro-1H-cyclopenta[a]phenanthren-17-yl)
pentanoate (43)
To a solution of 33 (42.9 mg, 0.14 mmol) and 38 (76.0 mg, 0.13 mmol) in CH
2
Cl
2
(1.3 mL) was added DMAP (1.6 mg, 0.013 mmol) and
DCC (28.9 mg, 0.14 mmol) at 0 C and the mixture was stirred at room temperature for 17.5 h. The crude solution was purified by flash
ꢀ
1
column chromatography (silica gel: 10 g, 70% AcOEt/hexane) to give 43 (39.3 mg, 34%). H NMR (300 MHz, CDCl
d, J = 8.6 Hz), 6.52 (1H, dd, J = 8.6, 2.4 Hz), 6.37 (1H, d, J = 2.4 Hz), 4.36-4.33 (2H, m), 4.23-4.20 (2H, m), 3.93-3.49 (20H, overlapped),
3
) d [ppm]: 7.42 (1H,
13
2
.43-2.17 (2H, m), 1.97-1.10 (27H, overlapped), 1.04 (9H, s), 0.91-0.89 (6H, overlapped), 0.64 (3H, s), 0.26 (6H, s). C NMR (75 MHz,
CDCl ) d [ppm]: 174.4, 163.0, 160.2, 154.4, 135.9, 107.6, 106.2, 104.8, 85.7, 83.7, 72.0, 70.8 (2carbons), 70.74, 70.72, 70.71, 70.70,
9.3, 69.0, 64.7, 63.5, 56.6, 56.1, 55.6, 42.8, 42.2, 40.5, 40.3, 36.6, 36.0, 35.5 (2 carbons), 34.7, 31.3, 31.0, 30.7, 28.3, 27.3, 26.5, 25.8
3
6
+
(
3 carbons), 24.3, 23.5, 20.9, 18.39, 18.35, 12.2, -4.3 (2 carbons). HRMS (ESI) caled. for C50H80NaO11Si [M + Na] 907.53676, found
9
07.53533.
2
-((3-(2-Hydroxy-4-methoxyphenyl)propioloyl)oxy)ethyl (R)-4-((3R,5R,8R,9S,10S,13R,14S,17R)-3-Hydroxy-10,13-
dimethylhexadecahydro-1H-cyclopenta[a]phenanthren-17-yl)pentanoate (7)
To a solution of 39 (37.5 mg, 0.053 mmol) in THF (5.3 mL) was added TBAF (0.11 mL, 0.11 mmol) at 0 C and the mixture was stirred for
h. The crude solution was purified by flash column chromatography (silica gel: 10 g, 60% AcOEt/hexane) to give 7 (18.2 mg, 58%).
ꢀ
1
1
H NMR (300 MHz, CDCl
3
) d [ppm]: 7.37 (1H, d, J = 8.5 Hz), 6.92 (1H, brs), 6.49-6.46 (2H, overlapped), 4.46-4.43 (2H, m), 4.36-4.33
13
(
2H, m), 3.81 (3H, s), 3.68-3.61 (1H, m), 2.53-2.22 (2H, m), 1.95-0.95 (27H, overlapped), 0.93-0.90 (6H, overlapped), 0.63 (3H, s).
C
Cell Chemical Biology 27, 1–16.e1–e11, March 19, 2020 e5