Abstract
Thymosin alpha 1 (Tα1) is an important immunomodulating agent with various clinical applications. The natural form of Tα1 is Nα -acetylated, which was supposed to be related to in vivo stability of the hormone. In this study, fusion protein Tα1-HSA was constructed and expressed in Pichia pastoris. RimJ, a Nα -acetyltransferase from E.coli, was also overexpressed and purified to homogeneity. In vitro acetylation of Tα1-HSA in the presence of RimJ and acetyl coenzyme A resulted in Nα -acetyl Tα1-HSA. The Nα -acetylation was determined by LC-MS/MS. Kinetic assay indicated that RimJ had a higher affinity to desacetyl Tα1 than to Tα1-HSA. Bioactivity assay revealed fully retained activity of Tα1 when the hormone was connected to the N-terminus of the fusion protein, while the activity was compromised in our previously constructed HSA-Tα1. With fully retained activity and N-terminal acetylation, Nα -acetyl Tα1-HSA was expected to be a more promising pharmaceutical agent than Tα1.