Recombinant Ganoderma lucidum immunomodulatory protein modified with polyethylene glycol

Recombinant Ganoderma lucidum immunomodu-latory protein (rLZ-8) expressed using the Pichia yeast eukaryotic expression system is a potential new drug for cancer therapy; however, it has a short half-life in the body. In order to optimize the potency and stability of rLZ-8, we modified the recombinant protein chemically using methoxy-PEG-succinimidyl propionate (mPEG-SPA). The results indicated that several parameters, including pH, the molar ratio of rLZ-8 to mPEG-SPA and time, played crucial roles in the modification process. In particular, when the molar ratio of rLZ-8 to mPEG-SPA was 1:1, rLZ-8 was modified by a single mPEG-SPA moiety. In addition, MALDI-TOF/TOF and ESI Q-Trap results revealed that the difference in molecular weight (MW) between the peptide-linked mPEG-SPA and the mPEG-SPA closely matched the MW of a methionine amino acid. Taken together, these data suggest that modification of mPEG-SPA occurred on the N-terminal helix of rLZ-8. This modification method has laid a foundation for the development of long-acting formulations of rLZ-8.