S-1

Synthesis of 2-deoxy-2,2-difluoro-α-maltosyl fluoride and its X-ray structure in complex with Streptomyces coelicolor GlgEI-

V279S Sandeep Thanna‡, Jared J. Lindenberger‡, Vishwanath V. Gaitonde, Donald R. Ronning* and

Steven J. Sucheck*

Department of Chemistry, The University of Toledo, 2801 W. Bancroft Street, MS602, Toledo, OH, USA 43606

Supplementary Information Page S-1: Contents

Page S-5: Enzyme substrate assay for α-MTF

Page S-6: Fig. S-1. Evaluation of α-MTF as a substrate for Sco GlgEI-V279S.

Page S-7: Synthesis of per-O-acetyl-2-fluoro-maltal (1) and 3,6-di-O-acetyl-4-O-(2’,3’,4’,6’-tetra-O-acetyl-α-D-glucopyranosyl)-2-deoxy-2-fluoro-α-D-mannosyl bromide (14).

Page S-8: Scheme S-1. Synthesis of per-O-acetyl-2-fluoro-maltal (1) and 3,6-di-O-acetyl-4-O-(2’,3’,4’,6’-tetra-O-acetyl-α-D-glucopyranosyl)-2-deoxy-2-fluoro-α-D-mannosyl bromide (14). Page S-9 to S-15: Experimentals for scheme S-1

2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-1,2,3,6-tetra-O-acetyl-α-D-glucopyranose (8)2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-2,3,6-tri-O-acetyl-α-D-glucopyranosyl bromide (9)2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-α-D-glycal (10)2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-α/β-D-gluco-and D-mannopyranoses (11)2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-1,3,6-tri-O-acetyl-2-deoxy-2-fluoro-α/β-D-gluco-and D-mannopyranoses (12)2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-α-D-gluco-and D-mannopyranosyl fluoride (13)2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-glycal (1)2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-α-D-mannopyranosyl bromide (14)

Page S-16: Synthesis 5-fluoro-β-D-idosyl fluoride (19) and 5-fluoro-α-glycosyl fluoride (20)

Page S-17: Scheme S-2

Electronic Supplementary Material (ESI) for Organic & Biomolecular Chemistry.This journal is © The Royal Society of Chemistry 2015

S-2

Page S-18: Fig. S-2. Reaction assembly set-up for photobromination of C5 position of glycosyl compounds

Page-S-19 to S-22: Experimentals for scheme S-21,2,3,4,6-penta-O-acetyl-α/β-D-glucopyranoside (15)2,3,4,6-penta-O-acetyl-α-D-glycosyl fluoride (16)2,3,4,6-penta-O-acetyl-5-bromo-α-D-glycosyl fluoride (17)2,3,4,6-O-acetyl-5-fluoro-β-D-idosyl fluoride (18)5-Fluoro-β-D-idosyl fluoride (19)5-Fluoro-α-D-glycosyl fluoride (20)

Pages S-23 to S-74: NMR Spectra

1H NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-2,3,6-tri-O-acetyl-α-D-glucopyranosyl bromide (9)13C NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-2,3,6-tri-O-acetyl-α-D-glucopyranosyl bromide (9)COSY of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-2,3,6-tri-O-acetyl-α-D-glucopyranosyl bromide (9)HMQC of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-2,3,6-tri-O-acetyl-α-D-glucopyranosyl bromide (9)1H NMR of 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-α-D-glycal (10)13C NMR of 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-α-D-glycal (10):COSY of 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-α-D-glycal (10)19F NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-α/β-D-gluco-and D-mannopyranoses (11)19F NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-1,3,6-tri-O-acetyl-2-deoxy-2-fluoro-α/β-D-gluco-and D-mannopyranoses (12)19 F NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-α-D-gluco/ D-mannopyranosyl bromide (13)1H NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-glycal (1) 13C NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-glycal (1)COSY of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-glycal (1) HMQC of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-glycal (1)19F NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-glycal (1) 1H NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-α-D-mannopyranosyl bromide (14)13C NMR attached protein test (APT) experiment of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-α-D-mannopyranosyl bromide (14)

S-3

COSY of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-α-D-mannopyranosyl bromide (14)HMQC of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-α-D-mannopyranosyl bromide (14)19F NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-α-D-mannopyranosyl bromide (14)1H NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-α-D-glucopyranosyl fluoride (2)13 C NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-α-D-glucopyranosyl fluoride (2)13C NMR (APT) of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-α-D-glucopyranosyl fluoride (2)COSY of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-α-D-glucopyranosyl fluoride (2)HMQC of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-α-D-glucopyranosyl fluoride (2)19F NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-α-D-glucopyranosyl fluoride (2) (Vxrs400 MHz)1H NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-β-D-glucopyranosyl fluoride (3) 13 C NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-β-D-glucopyranosyl fluoride (3)COSY of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-β-D-glucopyranosyl fluoride (3)HMQC of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-β-D-glucopyranosyl fluoride (3)19F NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-β-D-glucopyranosyl fluoride (3) (Gemini 200MHz)1H NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-α/β-D-glucopyranose (4) 13C NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-α/β-D-glucopyranose (4)COSY of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-α/β-D-glucopyranose (4)HMQC of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-α/β-D-glucopyranose (4)19F NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-α/β-D-glucopyranose (4)1H NMR of compound α-D-glucopyranosyl-(1,4)-2-deoxy-2,2-difluoro-α-D-glucopyranosyl fluoride (5)13C NMR of compound α-D-glucopyranosyl-(1,4)-2-deoxy-2,2-difluoro-α-D-glucopyranosyl fluoride (5)COSY of compound α-D-glucopyranosyl-(1,4)-2-deoxy-2,2-difluoro-α-D-glucopyranosyl fluoride (5)HMQC of compound α-D-glucopyranosyl-(1,4)-2-deoxy-2,2-difluoro-α-D-glucopyranosyl fluoride (5)

S-4

19F of compound α-D-glucopyranosyl-(1,4)-2-deoxy-2,2-difluoro-α-D-glucopyranosyl fluoride (5)1H NMR of compound α-D-glucopyranosyl-(1,4)-2-deoxy-2,2-difluoro-β-D-glucopyranosyl fluoride (6)13C NMR of compound α-D-glucopyranosyl-(1,4)-2-deoxy-2,2-difluoro-β-D-glucopyranosyl fluoride (6)COSY of compound α-D-glucopyranosyl-(1,4)-2-deoxy-2,2-difluoro-β-D-glucopyranosyl fluoride (6)HMQC of compound α-D-glucopyranosyl-(1,4)-2-deoxy-2,2-difluoro-β-D-glucopyranosyl fluoride (6)19F of compound α-D-glucopyranosyl-(1,4)-2-deoxy-2,2-difluoro-β-D-glucopyranosyl fluoride (6)1H NMR of compound 2,3,4,6-penta-O-acetyl-α-D-glycosyl fluoride (16)1H NMR of compound 2,3,4,6-penta-O-acetyl-5-bromo-α-D-glycosyl fluoride (17)1H NMR of compound 2,3,4,6-O-acetyl-5-fluoro-β-D-idosyl fluoride (18) 1H NMR of compound 5-fluoro-β-D-idosyl fluoride (19)19F-NMR of compound 5-fluoro-β-D-iodosyl fluoride (19) 1H-NMR of compound per-O-acetyl-5-fluoro-α-glycosyl fluoride (20)

Page S-75: References

S-5

Enzyme substrate assay for α-MTF

MALDI-MS was performed to evaluate α-MTF incorporation into maltooligosaccharides via Sco

GlgE-V279S and compared to a series of controls, (Fig S-1). The spectra of only maltohexaose

(M6) in buffer, showed a single peak representing a sodiated M6, Fig S-1A. The addition of Sco

GlgE1-V279S resulting in a series of higher molecular weight products which result from known

background transglycosylation activity catalyzed by Sco GlgE1-V279S, Fig S-1B. We also

prepared -maltosyl fluoride (-M1F), a substrate for Sco GlgE1. The use of M1F, M6 and Sco

GlgE1-V279S, Fig S-1C, resulted in higher molecular weight peaks. A final experiment replaced

M1F with α-MTF, which produced a spectrum identical to the control shown in Fig S-1B. Any

production of longer maltooligosaccharides would be the result of GlgE activity and those

products would have a molecular weight change resulting from the difluoromethylene unit.

S-6

Fig S-1. Evaluation of α-MTF as a substrate for Sco GlgEI-V279S. (A) control 1: ESI-MS of maltohexaose (M6). (B) control 2: ESI-MS of M6 + Sco GlgEI-V279S. (C) control 3: ESI-MS of M6 + Sco GlgEI-V279S + -M1F. (D) ESI-MS of M6 + Sco GlgEI-V279S + α-MTF. M4, M6, M8, M10, M12, and M14, represent 4, 6, 8, 10, 12, and 14 maltose units in the respective products. R = the continuation of the maltose repeat.

S-7

Synthesis of 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-

fluoro-glycal (1) and 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-

deoxy-2-fluoro-α-D-mannopyranosyl bromide (14)

Synthesis of Peracetylated glycal (10) was performed the methods of Braitsch et al.1 D-maltose 7

was peracetylated to obtain peracetyl maltose 8. Maltose derivative 8 was brominated at the

anomeric position using 33% hydrobromic acid in glacial acetic acid to obtain the corresponding

α-maltosyl bromide (9), Scheme S-1. Maltosyl bromide 9 was subjected to a zinc-mediated β-

elimination using activated zinc and N-methylimidazole to afford the glycal 10 in 53% yield.1

Glycal 10 was dissolved in DMF:H2O (3:1) and subjected to treatment with 1-(chloromethyl)-4-

fluoro-1,4-diazoniabicyclo[2.2.2]octane ditetrafluoroborate (Selectfluor®) to afford an anomeric

mixture of (α/β)-1-hydroxy-2-fluoro derivatives with gluco and manno (2:1 ratio, respectively)

stereochemistry 11.2 This mixture 11 was acetylated using Ac2O/Py. The resulting peracetylated

mixture 12 was treated with 33% hydrobromic acid in glacial acetic acid at 0 °C to afford a

mixture of 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-

α-D-gluco/ D-mannopyranosyl bromides (13). The mixture 13 was subjected to a second

elimination. Only the compound with gluco stereochemistry which has anti-periplanarity

between C1'-bromo group and the C2' hydrogen eliminated to afford per-O-acetyl-2-fluoro-

maltal (1).3, 4 This result was confirmed by isolating 2',3',4',6'-Tetra-O-acetyl-α-D-

glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-α-D-mannopyranosyl bromide (14)

while purifying glycal (1).

S-8

Scheme S-1. Synthesis of 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-glycal (1) and

2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-α-D-mannopyranosyl bromide (14).

O

RORO

RO

OR

O

ROROO

OR

OR

O

AcOAcO

AcO

OAc

O

AcOAcOO

OAc

Br

O

AcOAcO

AcO

OAc

OAcOO

OAc

O

AcOAcO

AcO

OAc

O

FAcOO

OAc

OR

11 R = H

12 R = Ac

O

AcOAcO

AcO

OAc

OAcOO

OAc

BrF

O

AcOAcO

AcO

OAc

OAcOO

OAc

F

b c

d b e

9 10

13

1

O

AcOAcO

AcO

OAc

OAcOO

OAc

Br

F

14

7 D-Maltose

8 R = Aca

a

Reagents and conditions: a) Ac2O, pyridine, rt (99%); b) 33% HBr in AcOH, dry CH2Cl2 (88%); c) Zinc, N-methylimidazole,

EtOAc, reflux (53%); d) Selectfluor®, DMF:H2O (3:1), rt (53%) (2:1); e) CH3CN, Et3N, reflux (30%).

S-9

2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-1,2,3,6-tetra-O-acetyl-α-D-

glucopyranose (8)1

To a solution of maltose monohydrate 7 (4.0 g, 11.1 mmol) in pyridine (12.5 mL, 133.2

mmol) was added acetic anhydride (14.3 mL, 178 mmol) drop wise at 0 C. The solution

was maintained at room temperature for 12 h. The pyridine was removed as an azeotrope

with toluene by repeated coevaporation. The resulting clear syrup was dissolved in ethyl

acetate and washed with 1 N HCl (25 mL) and saturated NaHCO3 solution (50.0 mL).

The organic layer was dried by passing through anhydrous Na2SO4. The solvent was

evaporated under vacuum to afford a clear white foam compound 8. Yield: 96% (7.2 g);

silica gel TLC Rf = 0.56 (1:1 hexane-acetone); 1H-NMR (600 MHz, CDCl3): δ 5.75 (d,

1H, 3J1,2 8.07 Hz, H-1), 5.42 (d, 1H, 3J1’,2’ 4.03 Hz, H-3'), 5.36 (dd, 1H, 3J3’,4’ 9.9 Hz,

3J3’,2’ 10.27 Hz, H-3’), 5.30 (dd, 1H, 3J3,2 9.17 Hz, 3J3,4 8.80 Hz, H-3), 5.06 (dd, 1H, 3J3’,4’

9.9 Hz, H-4'), 4.98 (dd, 1H, 3J2,3 8.80 Hz, H-2), 4.87 (dd, 1H, 3J2’,3’ 10.64 Hz, 3J3’,4’ 4.03

Hz, H-2'), 4.46 (dd, 1H, 2J6a,6b 12.10 Hz, 3J6a,5 1.83 Hz, H-6a), 4.26-4.22 (m, 2H, 6-a, 6-

b), 4.06-4.03 (m, 2H, 6’b, H-4), 3.95-3.93 (m, 1H, H-5’), 3.86-3.83 (m, 1H, H-5), 2.23 (s,

3H, CH3), 2.15 (s, 3H, CH3), 2.11 (s, 3H, CH3), 2.06 (s, 3H, CH3), 2.03 (s, 3H, CH3), 2.02

(s, 3H, CH3), 2.02 (s, 3H, CH3), 2.01 (s, 3H, CH3); 13C-NMR (600 MHz, CDCl3): 170.73

(C=O), 170.68 (C=O), 170.61 (C=O), 170.24 (C=O), 170.06 (C=O), 169.77 (C=O),

169.61 (C=O), 168.97 (C=O), 95.85 (C1'), 91.39 (C1), 75.40 (C3), 73.11 (C4), 72.50

(C5), 71.07 (C2), 70.13 (C2'), 69.43 (C3'), 68.71 (C5'), 68.06 (C4'), 62.64 (C6), 61.57

(C6'), 21.02 (CH3), 20.97 (2 CH3), 20.84 (CH3), 20.75 (3 CH3), 20.71 (CH3) ppm; mass

spectrum (ESIMS), m/z = 701.3 (M+23)+ C28H38O19 requires 701.19 (M+23)+.

S-10

2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-2,3,6-tri-O-acetyl-α-D-

glucopyranosyl bromide (9)1

Compound 8 (1.00 g, 1.47 mmol) was dissolved in dry methylene chloride (5.0 mL). To

the solution was added acetic acid (5.0 mL). The solution was cooled to 0 C 33% HBr in

acetic acid (1.0 mL, 2.76 mmol) was added by drop wise addition. The reaction was

maintained at 0 C for 4 h. Ice cold water was added and the solution extracted with DCM

(20.0 mL, 3 times). The combined DCM extracts were washed with cold distilled. H2O

and the organic layer dried over anhydrous Na2SO4. The solvent was removed under

reduced pressure to obtain a light yellow foam 9, which was sensitive to light. Yield:

88.2% (0.908 g); silica gel TLC Rf = 0.42 (6:4 Hexanes-Acetone). 1H-NMR (600 MHz,

CDCl3): δ 6.51 (d, 1H, 3J1,2 3.67 Hz, H-1), 5.63 (dd, 1H, 3J3,2 9.54 Hz, H-3), 5.44 (d, 1H,

3J1',2' 4.03 Hz, H-1'), 5.39 (dd, 1H, 3J3’,2’ 10.27 Hz, 3J3’,4’ 9.90 Hz, H-3), 5.09 (dd, 1H,

3J4’,3’ 9.90 Hz, H-4'), 4.88 (dd, 1H, 3J1’,2’ 4.03 Hz, 3J2’,3’ 10.64 Hz, H-2'), 4.73 (dd, 1H,

3J1,2 3.67 Hz, 3J2,3 9.90 Hz, H-2), 4.53 (dd, 1H, 2J6a,6b 13.94 Hz, 3J6a,5 4.55 Hz, H-6a),

4.29-4.25 (m, 3H, 6-a', 6-b, H-5), 4.10-4.05 (m, 2H, H-4, 6-b'), 3.96-3.95 (m, 1H, H-5'),

2.16 (s, 3H, CH3), 2.11 (s, 3H, CH3), 2.09 (s, 3H, CH3), 2.08 (s, 3H, CH3), 2.05 (s, 3H,

CH3), 2.04 (s, 3H, CH3), 2.02 (s, 3H, CH3) ppm; 13C NMR (CDCl3): 170.86 (C=O),

170.67 (C=O), 170.46 (C=O), 170.14 (2 C=O), 169.68 (C=O), 169.60 (C=O), 95.91 (C1'),

86.18 (C1), 72.66 (C5), 72.47 (C3), 71.64 (C4), 71.17 (C2), 70.15 (C2), 69.38 (C3'),

68.78 (C5), 68.01 (C4), 61.97 (C6), 61.45 (C6'), 21.03 (CH3), 20.95 (CH3), 20.86 (CH3),

20.81 (CH3), 20.77 (3 CH3) ppm; mass spectrum (ESIMS), m/z = 722.38 (M+23)+

C28H38O19 requires 722.44 (M+23)+.

S-11

2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-α-D-glycal (10)1

Compound 9 (2.10 g, 3.00 mmol) was dissolved in dry ethyl acetate (30 mL). To this

solution was added Zn powder (1.9 g, 30.0 mmol), and N-methylimidazole (377 mg, 4.60

mmol). The solution was stirred vigorously, heated to reflux, and maintained at that

temperature for 6 h. After the reaction was completed the Zn was removed by filtering

through Celite. The Celite was washed twice with EtOAc (15 mL) for complete

extraction. The filtrate was washed with saturated NaHSO4 (20 mL) followed by

NaHCO3 (20 mL), and the ethyl acetate layer was dried over anhydrous Na2SO4. The

dried ethyl acetate layer was concentrated under reduced pressure to obtain crude 10

which was purified by flash column chromatography on silica gel using a mobile phase of

25% acetone in hexanes. Compound 10 was obtained as a colorless solid. Yield: 53%

(0.891 g); silica gel TLC Rf = 0.44 (6:4 hexanes-acetone); 1H-NMR (600 MHz, CDCl3): δ

6.45 (d, 1H, 3J1,2 6.01 Hz, H-1), 5.52 (d, 1H, 3J1’,2’ 3.9 Hz, H-1'), 5.41 (dd, 1H, 3J3’,2’

10.01 Hz, H-3'), 5.18 (m, 1H, H-3), 5.07 (t, 1H, 3J3’,4’ 10.01 Hz, 3J4’,5’ 9.77 Hz, H-4'),

4.84-4.82 (m, 2H, H-2', H-2), 4.39 (dd,1H, 2J6a,6b 11.96 Hz, 3J6a,5 3.42 Hz, H-6a), 4.35

(dd, 1H, 2J6a,6b 12.21 Hz, 3J6a,5 5.62 Hz, H-6b), 4.31-4.29 (m, 1H, H-5), 4.25 (dd, 2J6’a,6’b

12.45, 3J6a,5 3.91 Hz, 1H, H-6'a), 4.10 (dd, 1H, 2J6’a,6’b 12.45 Hz, 3J6a,5 1.71 Hz, H-6'b),

4.06-4.02 (m, 2H, H-4, H-5'), 2.13 (s, 3H, CH3), 2.11 (s, 3H, CH3), 2.06 (s, 3H, CH3),

2.05 (s, 3H, CH3), 2.04 (s, 3H, CH3), 2.02 (s, 3H, CH3) ppm; 13C NMR (CDCl3): δ 170.8

(C=O), 170.71 (C=O), 170.71 (C=O), 170.61 (C=O), 170.26 (C=O), 170.78 (C=O),

145.78 (C1), 98.83 (C2), 95.97 (C1'), 74.25 (C5), 72.60 (C4), 70.59 (C2'), 69.826 (C3'),

69.82 (C3'), 69.74 (C3), 68.43 (C5'), 68.29 (C4), 62.03 (C6), 61.77 (C6'), 21.32 (CH3),

S-12

21.03 (CH3), 20.91 (CH3), 20.89 (CH3), 20.83 (CH3), 20.77 (CH3) ppm; mass spectrum

(ESIMS), m/z = 583.3 (M+23)+ C24H32O16 requires 583.49 (M+23)+.

2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-

α/β-D-gluco-and D-mannopyranoses (11)2

To a solution of compound 10 (1.22 g, 2.17 mmol) in DMF (15.0 mL) was added H2O

(5.0 mL). The reagent Selectfluor® (1.90 g, 5.41 mmol) was added to the solution in two

proportions. The second portion was added after 4 h of maintenance of the reaction at

room temperature. The reaction was allowed to stir an additional 12 h at room

temperature. The solution was diluted with EtOAc (60 mL) and washed with cold H2O

(50 mL 3 times). The organic layer was dried over anhydrous Na2SO4 and the solvent was

evaporated under reduced pressure to obtain a light yellow syrup. The crude product was

purified by flash column chromatography on silica gel by elution with acetone-hexanes

(2:3). This yielded a mixture of compounds, (α/β)-1-hydroxy-2-fluoro derivatives, with

gluco and manno stereochemistry 11. Yield: 54% (695.0 mg); silica gel TLC Rf = 0.31

(6:4 hexanes -acetone); crude product used for next step without purification; mass

spectrum (ESIMS), m/z = 619.25 (M+23)+ C24H33FO16 requires 619.16 (M+23)+.

2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-1,3,6-tri-O-acetyl-2-deoxy-2-

fluoro-α/β-D-gluco-and D-mannopyranoses (12)

To a solution compounds 11 (694 mg, 1.16 mmol) in pyridine was added acetic anhydride

(329 L, 3.49 mmol). The solution was maintained at room temperature for 8 h. The

reaction was concentrated by evaporating the pyridine under reduced pressure to obtain

S-13

an oily liquid. This residue was diluted with EtOAc (20 mL). The EtOAc layer was

washed with 0.5 N HCl (15 mL), followed by water (20 mL), and brine (20 mL). The

organic layer dried over anhydrous Na2SO4 and the solvent evaporated under reduced

pressure to obtain a colorless foam 12 as a mixture of α/β-2-fluoro derivatives with gluco

and manno stereochemistry 12 (2.2:1, respectively). Yield: 99% (742 mg); silica gel TLC

Rf = 0.48 (2:3 acetone-hexanes), the obtained compound was used in next step without

purification; 19F-NMR (Vxrs400 MHz, CDCl3): 2-fluro gluco derivative: δ -203.3 (dd,

2JF,2 48.84 Hz, JF,3 12.21 Hz, F-2a); 2-fluoro manno derivative: δ -204.75 (ddd, 2JF,2 48.83

Hz, 3JF,3 27.47 Hz, 3JF,1 7.64 Hz, F-2e) ppm; mass spectrum (ESIMS), m/z = 661.18

(M+23)+ C26H35FO17 requires 661.17 (M+23)+.

2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-

α-D-gluco-and D-mannopyranosyl bromide (13)

The mixture of compounds 12 (784 mg, 1.23 mmol) was dissolved in methylene chloride

(3.5 mL) and the solution was cooled to 0 C. To the solution was added 33% HBr in

acetic acid (4.5 mL) by drop-wise addition. The reaction was maintained at 18 C for 8 h.

After the reaction was complete, the solution was diluted with cold H2O and extracted

three times with DCM (25 mL). The organic layers were combined and washed with H2O

(15 mL), brine (15 mL), and dried over anhydrous Na2SO4. The solvent was evaporated

under reduced pressure to obtain a colorless foam 13 as a mixture of α-1-bromo-2-fluoro

derivatives with gluco and manno stereochemistry. Yield: 98% (807.0 mg); The crude

material was used for next step without purification; 19F-NMR of compounds α-bromo-2-

fluoro derivatives with gluco and manno stereochemistry: δ -183 (m, F-2), -190.5 (m, F-

S-14

2) ppm; mass spectrum (ESIMS), m/z = 681.09 (M+23)+ C24H32BrFO15 requires 681.08

(M+23)+.

2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-

glycal (1)4

Mixture 13 (808 mg, 1.23 mmol) was dissolved in acetonitrile (10.0 mL) and

triethylamine (304 µL, 3.68 mmol) was added. The reaction mixture was heated to 65-70

C for 6 h. The solution was concentrated under reduced pressure and purified by gravity

column chromatography on silica gel using a mobile phase consisting of 7% acetone in

DCM to obtain pure compound 1. Yield: 33% (230.0 mg); silica gel TLC Rf = 0.19 (3:97

acetone-dichloromethane), and compound 14. Yield: 30% (207.0 mg); silica gel TLC Rf

= 0.28 (3:97 acetone-dichloromethane); (1): 1H-NMR (600 MHz, CDCl3): δ 6.77 (d, 1H,

3J1,2 4.77 Hz, H-1), 5.44-5.41 (m, 2H, H-1', H-3'), 5.37 (dd, 1H, 3J3,4 5.87 Hz, 3J3,1 1.47

Hz, H-3), 5.05 (t, 1H, 3J3,4 9.9 Hz, H-4'), 4.85 (dd, 1H, 3J2',3' 10.27 Hz, 3J2',1' 4.03 Hz, H-

2'), 4.47 (m, 1H, H-5), 4.37 (dd, 1H, 2J6a,6b 11.74 Hz, 3J6a,5 7.70 Hz, H-6a), 4.20 (dd, 1H,

2J6'a,6'b 12.47 Hz, 3J6'a,5' 4.77 Hz, H-6a'), 4.16 (dd, 1H, 2J6a,6b 11.74 Hz, 3J6b,5 4.4 Hz, H-

6b), 4.13 (dd, 1H, 2J6'b,6'a 12.47 Hz, 3J6'b,5' 2.20 Hz, H-6b'), 4.09-4.06 (m, 1H, H-5), 4.00-

3.98 (m, 1H, H-4), 2.11 (s, 3H, CH3), 2.10 (s, 3H, CH3), 2.04 (s, 3H, CH3), 2.03 (s, 3H,

CH3) ppm; 13C-NMR (CDCl3): δ 170.59 (C=O), 170.47 (C=O), 170.31 (C=O), 170.68

(C=O), 169.97 (C=O), 169.59 (C=O), 142.12 (d, 1J2,2F 239.99 Hz, C2), 132.09 (d, 2J1,2F

39.61 Hz, C1), 96.88 (C1'), 74.50 (d, 3J4,2F 7.70 Hz, C4), 73.99 (C5), 70.54 (C2'), 69.87

(C3'), 68.38 (C4', C5'), 65.79 (d, 2J3,2F 23.11 Hz, C3), 61.91 (C6'), 60.62 (C6), 20.76 (s,

6H, 2 CH3), 20.71 (s, 3H, CH3), 20.68 (s, 3H, CH3), 20.62 (s, 3H, CH3), 20.55 (s, 3H,

S-15

CH3) ppm; 19F NMR (Vxrs400 MHz, CDCl3): δ -164.51 (dd ,1F, 3JF,1 6.11 Hz, 3J3,F 4.58

Hz, F-2) ppm; mass spectrum (ESIMS), m/z = 601.14 (M+23)+ C24H33FO16 requires

601.15 (M+23)+.

2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-

α-D-mannopyranosyl bromide (14)

1H-NMR (600 MHz, CDCl3): δ 6.37 (d, 1H, 3J1,2 9.54 Hz, H-1), 5.57 (d, 1H, 3J1',2' 4.04 Hz, H-1'),

5.54 (dd, 1H, 3J3,F 26.04 Hz, 3J3,4 9.54 Hz, 3J3,2 2.20 Hz, H-3), 5.40 (t, 1H, 3J3’4’ 9.90 Hz, H-3'),

5.10 (t, 1H, 3J3'4' 9.90 Hz, H-4'), 4.95 (d, 1H, 2JF,2 48.78 Hz, H-2), 4.89 (dd, 1H, 3J2’3’ 11.00 Hz,

3J2'1' 4.03 Hz, H-2'), 4.52 (dd, 1H, 2J6a,6b 12.47 Hz, 3J6a,5 1.83 Hz, H-6a), 4.32-4.29 (m, 2H, H-4,

H-6b), 4.27 (dd, 1H, 2J6'a,6'b 12.47 Hz, 3J6'a,5' 3.30 Hz, H-6a), 4.20-4.18 (m, 1H, H-5), 4.08 (dd,

1H, 2J6'a,6'b 12.47 Hz, 3J6'b,5' 2.20 Hz, H-6b'), 3.99-3.97 (m, 1H, H-5') ppm; 13C-NMR (CDCl3): δ

170.70 (C=O), 170.57 (C=O), 170.51 (C=O), 170.17 (C=O), 170.12 (C=O), 169.61 (C=O), 95.98

(C1'), 88.59 (dd, 1J2F,2 185.76 Hz, C2), 82.16 (d, 2J2F,1 26.14 Hz, C1), 72.98 (C5), 71.99 (C3),

71.88 (C2), 70.38 (C2'), 69.50 (C3'), 69.46 (C4), 68.74 (C5'), 68.03 (C4'), 62.03 (C5'), 61.49

(C6'), 21.07 (CH3), 20.91 (CH3), 20.84 (CH3), 20.78 (CH3), 20.75 (CH3), 20.67 (CH3) ppm; 19F

NMR: δ -183 (m, F-2 ) ppm; m/z = 681.0806 (M+Na)+, C6H10F2O5 requires 658.0909.

S-16

Synthesis of 5-fluoro-β-D-idosyl fluoride (19): 5-Fluoro-glycosyl fluorides have also been

reported to inhibit of glycoside hydrolases.5 We synthesized 5-fluoro-β-idosyl fluoride (19) and

trace 5-fluoro-α-glycosyl fluoride (20). 6, 7 We envisioned fluoride 19 as a key intermediate on

route to fluoride 20, Scheme S-2. The known approach to access C5' fluoro glycosides has been

to photo-brominate the C5' position of the glycoside followed by displacement of the bromide

using a fluoride source.8, 9 We explored the photo-bromination of per-O-acetyl-α-glycosyl

fluoride (16) to access 2,3,4,6-tetra-O-acetyl-5-bromo-1-fluoro-α-glycosyl fluoride (17) (Figure

S-2). Glycosyl fluoride 17 was treated with AgF or AgBF4; however, only undesired epimeric

2,3,4,6-tetra-O-acetyl-5-fluoro-β-L-idosyl fluoride 18 was formed.8-10 We investigated methods

for equilibrating 18 to the D-gluco configuration 20. We examined: i) longer reaction times, ii)

higher tempratures, and iii) addition of HF-pyridine to equilibriate the L-idosyl fluoride 18 to the

D-glucosyl fluoride 20.9 However, these efforts did not prove to be helpful in our case. Other

strategies to epimerize from the L-idose to the D-glucose configuration were to treat the L-idosyl

fluoride with Lewis acids e.g., i) BF3.OEt2, ii) BF3.OEt2 + TMSOTf, iii) BF3.OEt2 + HF-

pyridine, and iv) Cp2(Zr)Cl2 + AgOTf, respectively. 9, 11-13 BF3.OEt2 worked with trace yields in

our hands to produce the 2,3,4,6-tetra-O-acetyl-5-fluoro-α-glycosyl fluoride (20) in an equilbium

favoring starting material 18.6 Fluoride 18 was deacetylated with NH3-MeOH to afford 19. Since

we could only produce 20 in only trace ammounts and since the structure of the GlgE 2-deoxy-2-

fluoro-α-maltosyl fluoride (α-MDF) in complex with Sco GlgEI-E423A14 was reported, there

was no need to further pursue this target. Our reaction apparatus description and detailed

conditions to produce 17 may be helpful to other researchers.

S-17

Scheme S-2. Synthesis of 2,3,4,6-tetra-O-acetyl-5-fluoro-β-D-idosyl fluoride (19) and 2,3,4,6-

tetra-O-acetyl-5-fluoro-α-glycosyl fluoride (20)

O

18

AcO

OAc

F

OAc

OAcF

OAcOAcO

AcO

OAc

OAc

OAcOAcO

AcO

OAc

F

O

19

HO

OH

F

OH

OHF

15 16 17

OAcOAcO

AcO

OAc

FBr

a b

dc

20

e OAcOAcO

AcO

OAc

FF

Reagents and conditions: a) HF-Pyridine, 83 %; b) N-bromosuccinamide, AIBN, CCl4, 2x250W

flood lamp (50 %); c) AgF, CH3CN (68 %); d) NH3-MeOH, quantitative; e) BF3.OEt2 (low).

S-18

Figure S-2. Reaction assembly set-up for photobromination of C5 position of glycosyl compounds

S-19

1,2,3,4,6-penta-O-acetyl-α/β-D-glucopyranoside (15):15 Compound 15 (10.0 g, 0.055 mol),

was dissolved in pyridine (48.2 g, 0.61 mol) and acetic anhydride (62.2 g, 0.61 mol). DMAP (10

mg) was added to the reaction flask and reaction stirred for a period of 4 h. Completion of the

reaction was monitored by TLC using ethyl acetate-hexanes (3:7). The reaction was concentrated

under reduced pressure to obtain colorless residue. The residue was dissolved in toluene and

concentrated under reduced pressure to remove most of the pyridine and residual acetic acid

generated during the reaction. The residue was diluted with ethyl acetate and was washed with 1

N HCl followed by saturated aqueous sodium bicarbonate. The organic layer was dried over

anhydrous Na2SO4 and concentrated under reduced pressure. The crude material 15 was obtained

as white amorphous solid. Yield: 99% (21.0 g). The characterization data for compound 15

matches the previously reported data.

2,3,4,6-penta-O-acetyl-α-D-glycosyl fluoride (16)16 HF-pyridine (~70% hydrogen fluoride in

~30% pyridine) (15 mL) was added to compound 15 (2.0 g, 5.1 mmol) in a polypropylene tube.

The mixture was stirred to dissolution at room temperature. The reaction was further stirred for 7

h. Completion of the reaction was monitored by TLC using hexanes-acetone (6:4). The reaction

was diluted with excess of dichloromethane. The solution was neutralized with saturated aqueous

sodium bicarbonate cooled over an ice-bath. The organic layer was extracted with saturated

aqueous sodium bicarbonate solution (3 × 100 mL) to remove residual acid. The organic layer

was dried over anhydrous MgSO4 and concentrated under reduced pressure. The crude material

was purified using flash column chromatography on silica gel using 25% acetone in hexanes to

obtain α-glucosyl fluoride, compound 16 as white amorphous solid. Yield: 75% (1.32 g); silica

gel TLC Rf = 0.50 (99:2 ethyl acetate-dichloromethane). The characterization data for compound

S-20

16 matches the previously reported data; 1H NMR (600 MHz, CD3OD): δ (600 MHz, CDCl3)

5.76 (dd, 1H, 2JH1,F = 53.4 Hz, 3JH1,H2 = 2.4Hz, H-1), 5.49 (m, 1H, H-3), 5.15 (m, 1H, H-4), 4.95

(dddd, 1H, 3JH2,F = 24 Hz, 3JH2,H3 = 10.2 Hz, 3JH2,H1 = 2.4 Hz, H-2), 4.29 (dd, 1H, 3JH6a,H5 = 12.6

Hz, 3JH6a,H6b = 3.6 Hz, H-6a), 4.19 (m, 1H, H-5), 4.15 (dd, 1H, 3JH6b,H5 = 12.6 Hz, 3JH6a,H6b = 2.4

Hz, H-6b), 2.11 (s, 3H, CH3), 2.10 (s, 3H, CH3), 2.04 (s, 3H, CH3), 2.03 (s, 3H, CH3) ppm; mass

spectrum (ESIMS), m/z = 373.09 (M+23)+ C14H19FO9 requires 373.09 (M+23)+.

2,3,4,6-penta-O-acetyl-5-bromo-α-D-glycosyl fluoride (17)8 Compound 16 (1.80 g, 5.14

mmol) was stirred in 80.0 mL anhydrous CCl4 at room temperature, in a quartz flask attached to

a reflux assembly under nitrogen atmosphere. N-Bromosuccinimide (3.65 g, 20.6 mmol) and

AIBN (83 mg, 0.51 mmol) was added to the reaction vessel. The reaction was stirred under

refluxing conditions using 2x250 Watts (120V) heat lamps stationed at a distance of 3.0 cm on

either side of the reaction vessel. Completion of reaction was monitored using 1H-NMR and TLC

using ethyl acetate: dichloromethane (0.2:9.9). The reaction was stirred under reflux for 18.0 h.

The reaction was concentrated under reduced pressure to obtain a brown colored residue. The

residue was diluted with dichloromethane (100 mL) and washed sequentially with water (1 × 50

mL), saturated sodium bicarbonate (1 × 50 mL) and brine solution (1 × 50 mL). The organic

layer was dried over anhydrous MgSO4 and concentrated under reduced pressure. The crude

material was purified using silica gel flash column chromatography using 1 % ethyl acetate in

dichloromethane to obtain 5-bromo-α-glucosyl fluoride, compound 17. Yield: 47% (1.03 g);

silica gel TLC Rf = 0.55 (99:2 ethyl acetate-dichloromethane). The characterization data for

compound 17 matches the previously reported data. 1H NMR (600 MHz, CD3OD): δ 5.93 (dd,

1H, 2JH1,F = 52.8 Hz, 3JH1,H2 = 3.0 Hz, H-1), 5.86 (t, 1H, H-3), 5.21 (d, 1H, 3JH4,H3 = 10.2 Hz, H-

S-21

4), 5.06 (dddd, 1H, 3JH2,F = 24 Hz, 3JH2,H3 = 10.8 Hz, 3JH2,H1 = 3.6 Hz, H-2) 4.43 (dd, 2H, H-

6a,H-6b), 2.13 (s, 3H, CH3), 2.12 (s, 3H, CH3), 2.10 (s, 3H, CH3) 2.05 (s, 3H, CH3) ppm; mass

spectrum (ESIMS), m/z = 451.00 (M+23)+ C14H18BrFO9 requires 451.00 (M+23)+.

2,3,4,6-O-acetyl-5-fluoro-β-D-idosyl fluoride (18):8 Compound 17 (1.0 g, 2.2 mmol) was

dissolved in 15 mL of anhydrous acetonitrile under nitrogen atmosphere. The reaction flask was

covered with aluminum foil and the reaction carried out in the dark. Silver (I) fluoride (0.70 g,

5.5 mmol) was added to the flask under nitrogen atmosphere and the reaction stirred at room

temperature. The reaction was stirred for 4 h. Completion of the reaction was monitored by TLC

using ethyl acetate-hexane (3:97). The reaction was filtered and the filtrate was concentrated

under reduced pressure to obtain a light brown residue. The residue was dissolved in ethyl

acetate (50 mL) and washed sequentially with water (1 × 50 mL) and brine solution (1 × 50 mL).

The organic layer was dried over anhydrous MgSO4 and concentrated under reduced pressure.

The crude material was purified using silica gel flash column chromatography using 1 % ethyl

acetate in diethyl ether to obtain 5-fluoro-β-L-idosyl fluoride, compound 18. Yield: 68% (0.55

g); silica gel TLC Rf = 0.38 (3:97 ethyl acetate-hexanes). 1H NMR (600 MHz, CD3OD): δ 5.78

(d, 1H, 2JF,1H = 60.04, H-1), 5.39 (dd, 1H, 3JH2,F = 30.0 Hz, 3JH2,H3 = 12.0 Hz, H-2), 5.28 (m, 2H,

H-3, H-4), 4.43 (dd, 2H, 2JH6a,H6b = 24.0 Hz, 3JH6a,H5 = 12.0 Hz, H-6a), 4.18 (t, 1H, 2JH6a,H6b =

24.0 Hz, H-6b), 2.13 (s, 3H, CH3), 2.12 (s, 6H, 2CH3), 2.09 (s, 3H, CH3) ppm; mass spectrum

(ESIMS), m/z = 391.07 (M+23)+ C14H18F2O9 requires 391.08 (M+23)+.

5-Fluoro-β-D-idosyl fluoride (19):6 Compound 18 (8.6 mg) was dissolved in MeOH (1.5 mL)

and the reaction mixture was cooled to -10 C using dry ice in acetone. Ammonia gas was

S-22

bubbled into the solution for 10 min. The outlet of the reaction apparatus was connected to a bent

finger trap cooled to -78 C to trap any escaping ammonia gas. The reaction was maintained at

room temperature for 2 h. The reaction was concentrated to obtain an oily liquid of crude

compound 19, the crude compound was dissolved in minimum amount of H2O purified by C-18

chromatography (2% methanol in H2O), to obtain pure compound 19. 1H NMR (600 MHz,

CD3OD): δ 5.65 (d, 1H, 2JF,1H = 56.12, H-1), 3.93 (m, 1H, H-2), 3.78 (m, 3H), 3.66 (m, 1H) ppm;

mass spectrum (HRMS), m/z = 223.0401 (M+Na)+, C6H10F2O5 requires 223.0394 (M+Na)+.

5-Fluoro-α-D-glycosyl fluoride (20):17 Compound 19 (69.0 mg, 188 µM) was dissolved in

anhydrous CH2Cl2 (1.0 mL) under nitrogen atmosphere. The solution was cooled to 0 C and

BF3.OEt2 (0.007 mL) was added. The reaction was maintained at 0 C for another 10 minutes and

allowed to room temperature. The progress of reaction was monitored by 1H NMR. After 4 h

reaction was complete. The reaction was quenched with saturated aqueous NaHCO3 (5.0 mL).

The product was extraced from the aqueous layer by CH2Cl2 (5.0 mL). The organic layer was

washed with brine and dried over anhydrous MgSO4. The CH2Cl2 layer was concentrated under

reduced pressure and purifed by flash column chromatography on silica gel using 5% ethyl

acetate in dichloromethane as an eluent. The product was obtaind as mixture with 5-fluoro-β-D-

idosyl fluoride (19) and (20). 1H NMR (600 MHz, CD3OD): δ 5.85 (d, 1H, 2JF,1H = 53.0, H-1), 5.0

(dd, 1H, 3JH2,F = 24.0 Hz, H-2), 5.77 (t, 1H, 3JF,2H = 10.2, H-3), 5.23 (dd, 1H, 3JH4,F5 = 22.2 Hz,

H-4), 4.3 (dd, 1H, 2JH6a,H6b = 12.0 Hz, H-6a), 4.0 (dd, 1H, 2JH6a,H6b = 12.3 Hz, H-6b); mass

spectrum (ESIMS), m/z = 391.27 (M+Na)+, C6H10F2O5 requires 391.27 (M+Na)+.

S-23

1H NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-2,3,6-tri-O-acetyl-α-D-glucopyranosyl bromide (9):

9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0 -0.5Chemical Shift (ppm)

2.983.093.063.093.023.123.071.072.043.351.031.011.151.141.141.061.011.01

TMS2.02

2.04

2.05

2.08

2.09

2.11

2.16

3.95

4.07

4.08

4.25

4.25

4.27

4.29

4.53

4.54

4.72

4.74

4.88

4.89

4.89

5.09

5.37

5.39

5.43

5.44

5.63

6.51

6.52

7.27

S-24

13C NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-2,3,6-tri-O-acetyl-α-D-glucopyranosyl bromide (9)

S-25

COSY of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-2,3,6-tri-O-acetyl-α-D-glucopyranosyl bromide (9)

S-26

HMQC of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-2,3,6-tri-O-acetyl-α-D-glucopyranosyl bromide (9)

S-27

S-28

1H NMR of 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-α-D-glycal (10)

9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0 -0.5Chemical Shift (ppm)

3.073.046.073.043.103.151.041.072.012.011.040.940.980.950.94

TMS2.02

2.04

2.05

2.06

2.11

2.13

4.04

4.05

4.11

4.11

4.35

4.36

4.38

4.38

4.82

4.82

4.82

4.83

4.84

4.84

5.07

5.18

5.41

5.51

5.52

6.44

6.45

S-29

13C NMR of 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-α-D-glycal (10):

192 184 176 168 160 152 144 136 128 120 112 104 96 88 80 72 64 56 48 40 32 24 16 8 0Chemical Shift (ppm)

S-30

COSY of 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-α-D-glycal (10)

6.5 6.0 5.5 5.0 4.5 4.0 3.5F2 Chemical Shift (ppm)

4.0

4.5

5.0

5.5

6.0

6.5

F1 C

hem

ical

Shi

ft (p

pm)

S-31

19F NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-α/β-D-gluco-and D-mannopyranoses (11)

-80 -100 -120 -140 -160 -180 -200 -220 -240 -260 -280 -300 -320 -340Chemical Shift (ppm)

-205

.08

-204

.96

-204

.88

-204

.74

-200

.57

-200

.43

-200

.40

S-32

19F NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-1,3,6-tri-O-acetyl-2-deoxy-2-fluoro-α/β-D-gluco-and D-mannopyranoses (12)

-32 -40 -48 -56 -64 -72 -80 -88 -96 -104 -112 -120 -128 -136 -144 -152 -160 -168 -176 -184 -192 -200 -208 -216 -224 -232Chemical Shift (ppm)

-204

.84

-204

.71

-204

.64

-203

.38

-203

.25

-203

.23

-201

.63

-201

.53

S-33

19 F NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-α-D-gluco-and D-mannopyranosyl bromide (13)

20 10 0 -10 -20 -30 -40 -50 -60 -70 -80 -90 -100 -110 -120 -130 -140 -150 -160 -170 -180 -190 -200 -210 -220ppm

S-34

1H NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-glycal (1)

9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0 -0.5Chemical Shift (ppm)

3.003.032.828.771.081.213.281.051.040.991.021.022.100.95

2.03

2.04

2.10

2.11

2.11

4.00

4.14

4.14

4.16

4.17

4.19

4.20

4.37

4.39

4.84

4.84

4.85

4.86

5.05

5.06

5.41

5.42

5.42

5.44

6.77

6.77

S-35

13C NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-glycal (1)

190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0ppm

S-36

COSY of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-glycal (1)

S-37

HMQC of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-glycal (1)

S-38

19F NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-glycal (1)

0 -10 -20 -30 -40 -50 -60 -70 -80 -90 -100 -110 -120 -130 -140 -150 -160 -170 -180 -190 -200 -210 -220ppm

S-39

1H NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-α-D-mannopyranosyl bromide (14)

9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0 -0.5Chemical Shift (ppm)

2.993.032.993.002.953.041.021.061.023.201.031.540.501.031.020.521.531.00

2.03

2.04

2.06

2.11

2.14

2.16

2.18

2.20

3.99

4.09

4.26

4.26

4.29

4.31

4.31

4.32

4.51

4.52

4.88

4.89

4.90

4.91

5.08

5.10

5.38

5.40

5.56

5.57

6.37

6.39

7.27

S-40

13C NMR attached protein test (APT) experiment of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-α-D-mannopyranosyl bromide (14)

S-41

COSY of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-α-D-mannopyranosyl bromide (14)

S-42

HMQC of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-α-D-mannopyranosyl bromide (14)

S-43

19F NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2-fluoro-α-D-mannopyranosyl bromide (14)

20 10 0 -10 -20 -30 -40 -50 -60 -70 -80 -90 -100 -110 -120 -130 -140 -150 -160 -170 -180 -190 -200 -210 -220ppm

S-44

1H NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-α-D-glucopyranosyl fluoride (2)

9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0 -0.5Chemical Shift (ppm)

2.972.992.852.996.011.011.044.120.971.011.011.030.961.000.97

1.57

2.02

2.04

2.08

2.11

2.17

2.18

3.95

4.08

4.09

4.19

4.25

4.25

4.27

4.27

4.60

4.86

4.86

4.88

4.88

5.09

5.11

5.34

5.36

5.45

5.46

5.48

5.62

S-45

13 C NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-α-D-glucopyranosyl fluoride (2)

192 184 176 168 160 152 144 136 128 120 112 104 96 88 80 72 64 56 48 40 32 24 16 8 0 -8Chemical Shift (ppm)

101.

7210

1.96

102.

2010

3.26

103.

5010

3.74

112.

0011

2.24

113.

6611

3.90

115.

3411

5.59

S-46

13C NMR (APT) of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-α-D-glucopyranosyl fluoride (2)

S-47

COSY of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-α-D-glucopyranosyl fluoride (2)

S-48

HMQC of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-α-D-glucopyranosyl fluoride (2)

S-49

19F NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-α-D-glucopyranosyl fluoride (2) (Vxrs400 MHz)

8 0 -8 -16 -24 -32 -40 -48 -56 -64 -72 -80 -88 -96 -104 -112 -120 -128 -136 -144 -152 -160 -168 -176 -184 -192

Chemical Shift (ppm)

0.470.462.03

-14

8.3

8-1

48

.24

-12

2.6

1-1

22

.58

S-50

1H NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-β-D-glucopyranosyl fluoride (3)

9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0

Chemical Shift (ppm)

3.133.113.013.313.122.921.071.041.091.092.411.001.081.181.041.151.570.48

CHLOROFORM-d 2.03

2.04

2.07

2.11

2.15

2.18

3.97

4.10

4.10

4.20

4.23

4.25

4.25

4.27

4.84

4.85

4.86

4.86

5.08

5.10

5.38

5.39

5.42

5.43

5.44

S-51

13 C NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-β-D-glucopyranosyl fluoride (3)

190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0ppm

S-52

COSY of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-β-D-glucopyranosyl fluoride (3)

S-53

HMQC of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-β-D-glucopyranosyl fluoride (3)

S-54

19F NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro-β-D-glucopyranosyl fluoride (3) (Gemini 200MHz)

8 0 -8 -16 -24 -32 -40 -48 -56 -64 -72 -80 -88 -96 -104 -112 -120 -128 -136 -144 -152 -160 -168 -176 -184 -192

Chemical Shift (ppm)

1.011.001.01

-14

3.1

6-1

43

.09

-14

2.8

8

-12

8.6

2-1

27

.21

-11

5.5

0-1

14

.13

-11

4.0

8

S-55

1H NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro- α/β-D-glucopyranose (4)

9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0 -0.5Chemical Shift (ppm)

2.983.113.043.025.960.721.011.281.033.591.021.181.241.031.261.050.99

2.02

2.04

2.07

2.11

2.16

3.78

3.99

4.08

4.09

4.14

4.25

4.26

4.55

4.86

4.87

4.88

4.88

5.07

5.09

5.10

5.36

5.37

5.39

5.45

5.46

5.66

5.68

S-56

13C NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro- α/β-D-glucopyranose (4)

S-57

COSY of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro- α/β-D-glucopyranose (4)

S-58

HMQC of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro- α/β-D-glucopyranose (4)

S-59

19F NMR of compound 2',3',4',6'-Tetra-O-acetyl-α-D-glucopyranosyl-(1,4)-3,6-di-O-acetyl-2-deoxy-2,2-difluoro- α/β-D-glucopyranose (4)

0 -10 -20 -30 -40 -50 -60 -70 -80 -90 -100 -110 -120 -130 -140 -150 -160 -170 -180 -190 -200 -210Chemical Shift (ppm)

2.00

-182

.57

-182

.44

-182

.34

S-60

1H NMR of compound α-D-glucopyranosyl-(1,4)-2-deoxy-2,2-difluoro-α-D-glucopyranosyl fluoride (5)

9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0 -0.5Chemical Shift (ppm)

1.171.091.102.275.541.031.020.490.49

3.32

3.33

3.33

3.46

3.47

3.48

3.62

3.67

3.68

3.89

3.90

3.92

4.22

4.25

4.26

4.88

4.91

4.93

5.32

5.33

5.59

5.60

5.68

5.68

S-61

13C NMR of compound α-D-glucopyranosyl-(1,4)-2-deoxy-2,2-difluoro-α-D-glucopyranosyl fluoride (5)

192 184 176 168 160 152 144 136 128 120 112 104 96 88 80 72 64 56 48 40 32 24 16 8 0Chemical Shift (ppm)

S-62

COSY of compound α-D-glucopyranosyl-(1,4)-2-deoxy-2,2-difluoro-α-D-glucopyranosyl fluoride (5)

S-63

HMQC of compound α-D-glucopyranosyl-(1,4)-2-deoxy-2,2-difluoro-α-D-glucopyranosyl fluoride (5)

S-64

19F of compound α-D-glucopyranosyl-(1,4)-2-deoxy-2,2-difluoro-α-D-glucopyranosyl fluoride (5)

20 0 -20 -40 -60 -80 -100 -120 -140 -160 -180 -200 -220 -240Chemical Shift (ppm)

S-65

1H NMR of compound α-D-glucopyranosyl-(1,4)-2-deoxy-2,2-difluoro-β-D-glucopyranosyl fluoride (6)

9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0Chemical Shift (ppm)

1.091.051.053.043.121.021.051.000.500.49

3.29

3.29

3.30

3.30

3.33

3.57

3.63

3.63

3.64

3.65

3.82

3.84

4.04

4.09

4.87

4.90

5.26

5.27

5.40

5.42

5.49

5.51

S-66

13C NMR of compound α-D-glucopyranosyl-(1,4)-2-deoxy-2,2-difluoro-β-D-glucopyranosyl fluoride (6)

220 210 200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 -10 -20Chemical Shift (ppm)

S-67

COSY of compound α-D-glucopyranosyl-(1,4)-2-deoxy-2,2-difluoro-β-D-glucopyranosyl fluoride (6)

S-68

HMQC of compound α-D-glucopyranosyl-(1,4)-2-deoxy-2,2-difluoro-β-D-glucopyranosyl fluoride (6)

S-69

19F of compound α-D-glucopyranosyl-(1,4)-2-deoxy-2,2-difluoro-β-D-glucopyranosyl fluoride (6)

20 0 -20 -40 -60 -80 -100 -120 -140 -160 -180 -200 -220 -240Chemical Shift (ppm)

S-70

1H NMR of compound 2,3,4,6-penta-O-acetyl-α-D-glycosyl fluoride (16)

9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0Chemical Shift (ppm)

3.062.875.962.071.041.010.991.000.520.51

TMS2.03

2.05

2.11

4.14

4.14

4.16

4.16

4.20

4.28

4.29

4.30

4.31

4.95

5.14

5.16

5.17

5.48

5.50

5.52

5.71

5.71

5.80

5.80

S-71

1H NMR of compound 2,3,4,6-penta-O-acetyl-5-bromo-α-D-glycosyl fluoride (17)

9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0 -0.5Chemical Shift (ppm)

3.032.953.043.102.021.061.011.470.44

2.06

2.11

2.12

2.14

4.36

4.38

4.42

4.44

5.04

5.05

5.06

5.08

5.09

5.10

5.21

5.23

5.85

5.87

5.88

5.89

5.97

5.97

S-72

1H NMR of compound 2,3,4,6-O-acetyl-5-fluoro-β-D-idosyl fluoride (18)

9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0 -0.5Chemical Shift (ppm)

2.922.836.002.091.051.031.000.980.510.50

TMS2.03

2.05

2.11

2.11

4.14

4.16

4.16

4.18

4.20

4.28

4.29

4.30

4.31

4.95

5.14

5.16

5.17

5.48

5.50

5.52

5.71

5.71

5.80

5.80

S-73

1H NMR of compound 5-fluoro-β-D-idosyl fluoride (19)

9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0 -0.5Chemical Shift (ppm)

1.002.800.970.430.42

3.31

3.31

3.66

3.67

3.69

3.79

3.80

3.81

3.90

3.93

3.93

3.96

4.70

5.60

5.70

S-74

19F-NMR of compound 5-fluoro-β-D-iodosyl fluoride (19)

0 -10 -20 -30 -40 -50 -60 -70 -80 -90 -100 -110 -120 -130 -140 -150 -160 -170 -180 -190 -200 -210Chemical Shift (ppm)

1.011.00

-139

.64

-139

.58

-139

.51

-139

.35

-139

.21

-139

.14

-112

.69

-112

.59

-112

.54

S-75

1H-NMR of compound per-O-acetyl-5-fluoro-α-glycosyl fluoride (20)

9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0 -0.5Chemical Shift (ppm)

3.052.913.373.920.761.010.731.090.831.00

TMS2.05

2.12

2.13

2.14

4.07

4.07

4.09

4.09

4.30

4.31

4.32

4.33

5.04

5.06

5.10

5.30

5.31

5.33

5.35

5.76

5.78

5.80

5.81

5.81

5.89

5.90

S-76

References1. M. Braitsch, H. Kahlig, G. Kontaxis, M. Fischer, T. Kawada, R. Konrat and W. Schmid,

Beilstein J. Org. chemistry, 2012, 8, 448-455.2. M. D. Burkart, Z. Zhang, S. Hung and C. Wong, J. Am. Chem. Soc., 1997, 119, 11743-

11746.3. E. Boyd, R. V. H. Jones, P. Quayle and A. J. Waring, Tetrahedron Lett., 2006, 47, 7983-

7986.4. R. Zhang, J. D. McCarter, C. Braun, W. Yeung, G. D. Brayer and S. G. Withers, J. Org.

Chem., 2008, 73, 3070-3077.5. H. J. Rozeboom, S. Yu, S. Madrid, K. H. Kalk, R. Zhang and B. W. Dijkstra, J. Biol.

Chem., 2013, 288, 26764-26774.6. J. D. McCarter and S. G. Withers, J. Biol. Chem., 1996, 271, 6889-6894.7. R. Zhang, C. Li, L. K. Williams, B. P. Rempel, G. D. Brayer and S. G. Withers,

Biochemistry, 2009, 48, 10752-10764.8. B. W. Skelton, R. V. Stick, K. A. Stubbs, A. G. Watts and A. H. White, Aust. J. Chem. ,

2004, 57, 345-353.9. J. D. McCarter and S. G. Withers, J. Am. Chem. Soc., 1996, 118, 241-242.10. K. Igarashi, T. Honma and J. Irisawa, Carbohydr. Res., 1970, 13, 49-55.11. E. L. Myers, C. P. Butts and V. K. Aggarwal, Chem. Commun., 2006, 4434-4436.12. N. Chakraborty and M. d’Alarcao, Bioorg. Med. Chem., 2005, 13, 6732-6741.13. W. Bröder and H. Kunz, Bioorg. Med. Chem., 1997, 5, 1-19.14. K. Syson, C. E. Stevenson, A. M. Rashid, G. Saalbach, M. Tang, A. Tuukkanen, D. I.

Svergun, S. G. Withers, D. M. Lawson and S. Bornemann, Biochemistry, 2014, 53, 2494-2504.

15. A. Jha and N. Jain, Tetrahedron Lett., 2013, 54, 4738-4741.16. S. Dedola, D. L. Hughes and R. A. Field, Acta Crystallogr., Sect. C: Cryst. Struct.

Commun., 2010, 66, 124-127.17. J. D. McCarter and S. G. Withers, J. Am. Chem. Soc., 1996, 118, 241-242.