Practical Protein Chemistry - A. Darbre 1989

Determination of the composition of protein oligomers. Preparation of monomers and polypeptide chains
Preparation of monomers for sequencing
Cleavage of oligosaccharide fragments from glycoproteins

To cleave oligosaccharide fragments from Glycoproteins, a commercially available solution of HF in pyridine can be used (the reaction is carried out at room Temperature for 90 min). However, this reagent is not always as effective as anhydrous HF, which cleaves the glycosidic bonds of neutral and acidic sugars at 0 °C for 1 h [124]. Unfortunately, this Treatment alters the Physical Properties of the protein, although the sample may still be entirely suitable for sequencing.

1.6.4.1. Procedure [124]. The reaction with anhydrous hydrogen fluoride is performed in specialized apparatus, such as that supplied by Peninsula Lab. Inc. A thoroughly dried glycoprotein sample (up to 100 mg) and anhydrous methanol (up to 10% of the sample volume) are placed into the Reactor. The HF supply line is evacuated, with the exception of the liquid HF cylinder. The reactor is cooled with a dry ice-acetone bath, and 10 ml of liquid HF is distilled into it (cobalt trifluoride is used as a moisture scavenger in the liquid HF). Distillation is carried out while gently swirling both vessels. The reactor is allowed to warm naturally to the desired temperature (the vapor pressure of HF is 364 mmHg at 0 °C and 850 mmHg at 23 °C), and the reaction time is measured from this point onward. If the reaction is conducted at 0 °C, the reactor is cooled in an ice bath. Finally, the reactor is evacuated through a CaO trap. To prevent foaming of the reaction mixture, the vacuum should be applied gradually yet vigorously enough to rapidly drive off HF without causing oligomerization. After evaporating the HF, the line is kept under vacuum for an additional 1 h. Complete removal of HF is indicated by a color change of the reaction mixture from light brown or reddish to colorless. The sample is then dissolved in 0.1 M or 50% acetic acid and immediately chromatographed on a Sephadex Column or dialyzed.

Under more rigorous treatment with anhydrous HF (23 °C, 3 h), O-glycosidic bonds of amino sugars are cleaved, whereas peptide and N-glycosidic bonds remain unaffected.