Characterization of Three Species of Escherichia coli -Derived Human Leukocyte Interferon A Separated by Reverse-Phase High-Performance Liquid Chromatography

Abstract

A preparation of recombinant human leukocyte interferon A (rIFN-αA ) obtained from Escherichia coli cells was found by reverse-phase high-performance liquid Chromatography (HPLC) to contain three species. The three species, named Mf-1, Mf-2, and Ms in the order of their elution, were separated and characterized to elucidate their structural differences. The amino acid compositions, the amino-terminal amino acid sequences (Cys¹-Asp²-Leu³-), and the carboxy-terminal amino acids (Glu) of the three species agreed with those predicted from the cDNA sequence, although the sequence analysis yields of Mf-2 and Ms were extremely low. They were found to have the same disulfide bonds, Cys¹—Cys⁹⁸ and Cys²⁹—Cys¹³⁸, by amino acid analysis of the tryptic peptides, but the recovery of the peptides linked by a Cys¹—Cys⁹⁸ disulfide bond for Mf-2 and Ms was extremely low. The results demonstrate that although the primary structures of the three species are almost identical, small difference(s) exists among them especially at the amino-terminal portion.