PURIFICATION OF SYNTHETIC PEPTIDES WITH THE AID OF REVERSIBLE CHROMATOGRAPHIC PROBES

The application of reversible chromatographic probes for the purificat ion of synthetic peptides by reversed-phase (RP) and affinity chromato graphy is described. In previous work the concept of a one-step purifi cation procedure based on the acid-stable fluorenylmethoxycarbonyl (Fm oc) group was considered. Similar probes were used to purify peptides from 17 to 104 residues in length, using ion-exchange and RP media. Th e RP probes described here represent simplified versions of those repo rted previously. Further, the application of a new probe containing bi otin, for affinity chromatography, is demonstrated. Stepwise solid-pha se peptide synthesis was performed using both Boc and Fmoc chemistries and N-(2-chlorobenzyloxycarbonyloxy)succinimide as a capping reagent. The chromatographic probes were incorporated on to the ''target'' pep tide as active succinimidyl carbonate derivatives. After cleavage usin g either hydrogen fluoride or trifluoroacetic acid, purification was p erformed on either RP media or immobilized avidin. Treatment with orga nic base yielded the free purified product. The efficiency of the new probe molecules and the capping procedure was demonstrated by the puri fication of two polypeptides, 46 and 101 residues in length.