Work

Improving the bioactivity of rHirudin with boronophenylalanine site-specific modification

Year: 2015

Type: article

Abstract: To improve the bioactivity of recombinant (r)Hirudin, the orthogonal pair MjBTyrRS/tRNATyr cua (made up of the boronophenylalanine, tRNA and tRNA synthetase), was selected to incorporate boronophenylalanine site‑specifically into rHirudin at the 63 sites in an Escherichia coli system in response to the TAG codon. Following fusion with the gIII signal peptide and a hexahistidine tag, the modified protein was secreted into Luria‑Bertani culture medium and purified by nickel-nitrilotriacetic acid affinity chromatography following a gel filtration column. In a 200 ml flask, the yield of boronophenylalanine‑modified hirudin was 10 mg l‑1 and that of rHirudin was 19 mg l‑1. The authenticity of the purified proteins was verified using matrix-assisted laser desorption ionization time of flight mass spectroscopy and antithrombin activity assays. The results revealed that the antithrombin activity of the boronophenylalanine‑modified hirudin to human thrombin was more enhanced than that of rHirudin. The modified hirudin demonstrated stronger proliferation inhibiting ability on fibroblast L929 cells compared with that of rHirudin.

Source: Molecular Medicine Reports

DOI: 10.3892/mmr.2015.3194

Authors: XIUJUAN XIN, WENSHE LIU, LIYUN SUN

Institutions: East China University of Science and Technology, State Key Laboratory of Bioreactor Engineering, Texas A&M University

Language: English

Topic: Leech Biology and Applications

Subfield: Pharmacology

Field: Medicine

Domain: Health Sciences

SDG: Good health and well-being

Open Access status: bronze

Funders: State Key Laboratory of Bioreactor Engineering, Fundamental Research Funds for the Central Universities