0-glycosyl threonine

To Fmoc 0-glycosyl threonine (erf-butyl ester 20 (290 mg, 0.4 mmol) dissolved in dry dichloromethane (5 mL) is added dry trifluoroacetic acid (3 mL) at 0°C. The mixture is allowed to warm to room temperature, and the reaction is monitored by TLC (dichloro-methane-ethanol 10 1). After 3 h, the conversion is complete. The solvent is evaporated in vacuo. Toluene (10 mL) is codistilled in vacuo from the remainder. Small amounts of an unpolar impurity are separated by flash chromatography on silica gel (20 g) in dichloro-methane-ethanol (15 1) yield 230 mg (90%) [a]D 69.7° (c 1, CHC13) Rf 0.32 (CH -ethanol 10 1). 

Such electronic effects are readily deduced from chemical shifts. A recent example from our laboratory concerns the antimicrobial peptide drosocin.61 This 19-amino-acid peptide is secreted by Drosophila species in response to septic injury and is a potent antibacterial. The sequence of the peptide (shown below using the one-letter amino acid code) contains a glycosylated threonine at a midchain position that appears to be important in enhancing antimicrobial activity. 

Mann K, MechUng DE, Bachinger HP, Eckerskorn C, Gaill F, Timpl, R. Glycosylated threonine but not 4-hydroxyproline dominates the triple helix stabilizing positions in the sequence of a hydrothermal vent worm cuticle collagen. J. Mol. Biol. 1996 261 255-266. 

Af-(D-Biotinyl)-0-(3,4,6-tri-0-acetyl-2-azido-2-deoxy-a-D-galactopyranosyl)-L-threonine te/t-Butyl Ester 23 [34], A mixture of D-biotin (150 mg, 0.6 mmol), l-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDC 580 mg, 3 mmol), and 1-hydroxy-benzotriazole (HOBt 540 mg, 4 mmol) in dimethylformamide (DMF 2 mL) is stirred under exclusion of moisture at 22 C. After 45 min, the biotin is dissolved, and a solution of freshly prepared glycosyl threonine ester 22 (0.3 mmol, preceding procedure) in dichloro-methane (2 mL) is added at 0°C. After stirring for 16 h at room temperature, the solvent is evaporated in vacuo, the remainder dissolved in dichloromethane (50 mL), extracted with ice-cold 0.2 N HCl (3 x 25 mL), water (25 mL), and saturated NaHCOj solution (2 x 25 mL), dried with MgSO, and concentrated in vacuo. Purification by flash chromatography on silica gel (20 g) in dichloromethane-ethanol (25 1) yields 23 200 mg (93%) [a] 96.5° (c 1, CHClj) Rf 0.29 (toluene-acetone 4 1). 

Specifically, glycosylated threonine derivatives have been extended by solid phase peptide methodology to glycosylated hexapeptides related to oncofetal fibronectin the sugars involved were jS-D-GlcNAc and the (1-3)-linked y9-D-Gal derivative of it. "... 

This active ester was used for carboxyl protection of Fmoc-serine and Fmoc-threonine during glycosylation. The esters are then used as active esters in peptide synthesis. 

Human interleukin 2, a 133-residue protein, has been separated into multiple molecular forms by selective immunoaffinity chromatography and chromatofocusing. Most of the heterogeneity has been attributed to variations in glycosylation of the threonine residue in position 3 of the polypep-... 

C-N.m.r. Chemical-shift Data for the Anomeric-Carbon Atoms and C, and Coupling Constants ( /ch) Measured for Selected Gland /S-D-Glycosyl-L-threonine Model Compounds19- 1,82-84... 

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