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Updated: Jun 28, 2026
Surface Passivation for Single-molecule Protein Studies
Published on: April 24, 2014
[A comparative study on the properties of trichosanthin before and after site-directed PEGylation]
Qun-xing An1, Ying-feng Lei, Shi-jie Mu
1Department of Blood Transfusion, Xijing Hospital, Fourth Military Medical University, Xi'an 710032, China.
Objective:
To construct PEGylated trichosanthin (TCS) mutein and analyze its bioactivities, immunogenicity, acute toxicity, and pharmacokinetics.
Methods:
The potential antigenic determinant site YFF81-83 in the molecule of TCS was selected to undergo site-directed mutagenesis. Thus, a TCS mutein named TCS(YFF81-83ACS) was constructed and expressed in Escherichia coli of the line BL21 (DE3). Wild TCS (wTCS), TCSY(FF81-83ACS), and PEGylated TCS(YFF81-83ACS) (PEG- TCS(YFF81-83ACS)) of different concentrations were incubated with the supercoiled plasmid pUC19 to detect the DNAse activity, mixed with rabbit reticulocyte lysate to detect the ribosome inactivation activity, subcutaneously injected into 6 mice respectively to measure the serum IgG and IgE levels, intravenously injected into mice to observe the toxicity, and intravenously injected into SD rats to observe its -plasma half-life.
Results:
The DNAse activity of the PEG-TCS(YFF81-83ACS) was similar to that of the wTCS. The ribosome inactivation activity of the PEG-TCS(YFF81-83ACS) was 1/9-1/8 of that of the wTCS (P < 0.05). The serum IgE and IgG levels of the PEG-TCS(YFF81-83ACS) were both significantly lower than those of the wTCS (both P < 0.05). The LD50 of the PEG-TCS(YFF81-83ACS) was 1.8 times that of the wTCS (P < 0.05). The mean residence time and plasma half-life of the PEG-TCS(YFF81-83ACS) were significantly increased and its plasma clearance was significantly decreased (all P < 0.05).
Conclusion:
Site-directed mutagenesis and PEGylation of TCS provide a new approach for reconstructing TCS.
