Purification of the antibacterial fragments of guinea-pig major basic protein

Abstract

In this study, we tried to purify the antibacterial fragments of guinea-pig major basic protein (MBP) using Staphylococcus aureus. The antibacterial activity of MBP was not affected by the pyridylethylation, suggesting that the disulfide bonds were not necessary for the antibacterial activity. When pyridylethylated-MBP was digested with α-chymotrypsin, the four potent antibacterial fragments (fragment V (Arg 105-Tyr 119), fragment IX (Thr 1-Phe 67), fragment X (Ile 54-Leu 97) and fragment XIII (Arg 25-Phe 67)) were isolated by reverse-phase high-performance liquid chromatography. Anti-MBP monoclonal antibody, BMK-13 neutralized the antibacterial activity of PE-MBP and fragments IX, X and XII, but not the activity of fragment V, suggesting that Ile 54-Phe 67, the common amino-acid sequence of fragments IX, X and XIII, might be involved in the antibacterial activity of MBP. In fact, the synthetic peptide, Ile 54-Phe 67 exerted the antibacterial activity, and the activity was neutralized with BMK-13. The antibacterial activity of Ile 54-Phe 67 was lost by the modification with peptidylarginine deiminase which converted arginine residue to citrulline residue, suggesting that the arginine residues may be important for the antibacterial activity.