Binding of proline- and hydroxyproline-containing peptides and proteins tothe capillary wall

Sticking of peptides (as demonstrated by peak distortion) containing a high proportion of glycine and proline residues to the capillary wall was inves tigated. At acid pH where the carboxy terminal proline (e.g. Gly-Pro) is pr otonated, distinct sticking of these peptides occurred. On the contrary, th e Pro-Gly peptide yielded a well shaped peak. At alkaline pH, where the sit uation was reversed, the peptide sticking most to the wall was Gly-Gly, whi le the behaviour of the dipeptides possessing N- or C-terminal proline did not suffer from sticking to the wall. It was concluded that the separation of simple proline- and glycine-contain ing dipeptides can be partly optimized by manipulating the pH of the backgr ound electrolyte; particularly if cetyltrimethylammonium bromide, methanol or acetonitrile is added to the run buffer (reversed polarity mode with cet yltrimethylammonium bromide). However, all attempts to resolve more complex mixtures of proline- (and hydroxyproline-) containing di- and tripeptides failed; addition of an organic modifier, i.e. methanol and acetonitrile, he xylamine, triethylamine and cetyltrimethylammonium bromide was unsuccessful . Such separations can be materialized by using simple 25 mmol/l phosphate buffer at pH 10.5 provided that the sample is dissolved in (aqueous) 17.5 m mol/l Brij or 33 mmol/l sodium dodecyl sulfate. These systems are also appl icable to large polypeptides rich in proline, hydroxyproline and glycine: t ypically parent collagen alpha-chains, their dimers, trimers and higher pol ymers were successfully separated. (C) 1999 Elsevier Science B.V. All right s reserved.