PURIFICATION AND CHARACTERIZATION OF A HIGH-MOLECULAR-WEIGHT FORM OF RECOMBINANT HUMAN INTERLEUKIN-2

During purification of recombinant Interleukin-2 (rIL-2) by reversed-p hase HPLC, early fractions are discarded due to the presence of an uni dentified form of rIL-2. A procedure has been developed to isolate and purify this unidentified form of rIL-2. The purification process invo lves two chromatography steps and utilizes a Bakerbond Carboxy-Sulfon (CS) column under two different conditions. This material, designated as a high-molecular-weight form of rIL-2 (HMWrIL-2), exhibits lower mo bility during SDS-PAGE and has a pI which is approximately one unit le ss than that of rIL-2, but has similar bioactivity to rIL-2. Structura l analysis through enzymatic cleavage, HPLC peptide mapping, mass spec trometry, sequencing, and amino acid composition revealed that the dif ference between these two proteins is a C-terminal extension of 11 ami no acids. This extension could be the result of a nonstandard translat ion event.