Production of Nα-acetyl Tα1-HSA through in vitro acetylation by RimJ
Metrics: PDF 411 views | HTML 846 views | ?
Jing Chen1,*, Haibin Li1,*, Tao Wang2,*, Shuyang Sun3, Jia Liu1 and Jianhua Chen1
1School of Life Science and Technology, China Pharmaceutical University, Nanjing 210009, China
2Department of Neurosurgery, Shanghai 5th People’s Hospital, Shanghai Medical College, Fudan University, Shanghai 200240, China
3Overseas Education College, Nanjing Tech University, Nanjing 211816, China
*These authors have contributed equally to this work
Jianhua Chen, email: firstname.lastname@example.org
Keywords: thymosin alpha 1, human serum albumin, RimJ, acetylation, bioactivity
Received: May 30, 2017 Accepted: July 13, 2017 Published: August 14, 2017
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.
All site content, except where otherwise noted, is licensed under a Creative Commons Attribution 3.0 License.