Expression, Purification
and Preliminary Clinical Use of Recombinant HBsAg GST-PreS1(21–47 aa) Fusion
Proteins
WEI Jun, LIU Xiao-Jin, LI Guang-Di, WANG Yuan, ZHANG
Zu-Chuan
( Institute of Biochemistry and Cell Biology, Shanghai Institutes for
Biological Sciences, the Chinese Academy of Sciences, Shanghai 200031,
China )
WANG Yu-Qin, LU Zhi-Meng
( Rui Jin Hospital, Shanghai Second Medical University, Shanghai, 200025,
China )
Abstract Expression
plasmids pGEXSI and pGEXSII containing one copy and two orderly joined copies
of PreS1(21–47 aa) DNA fragment, respectively, were constructed.
GST-PreS1(21–47 aa) and GST-2×PreS1(21–47 aa) fusion proteins were highly
expressed in E.Coli TG1, induced by IPTG. The expression level of
GST-PreS1(21–47 aa) was about 30% of total soluble proteins in the lysate of
expression bacteria, and GST-2×PreS1(21–47 aa) was about 15% of total soluble
proteins, asestimated by SDS-PAGE. 50 mg GST-PreS1(21–47 aa) or 20 mg
GST-2×PreS1(21–47 aa) with purity over 90% was obtained, respectively, from 1
L culture by using affinity chromatography of glutathione Sepharose 4B. Direct
ELISA results showed that antigenicity of GST-2×PreS1(21–47 aa) was better
than GST-PreS1(21–47 aa) and synthetic peptide. Using GST-2×PreS1(21–47 aa)
as coated antigen, a sensitive indirect ELISA for detection of
anti-PreS1(21–47 aa) antibody, based on protein A-biotin and streptavidin HRP,
was established. The results from 99 sera samples of hepatitis B patients
showed that anti-PreS1(21–47 aa) antibody was detected in nearly half of acute
hepatitis B patients during recovery, but it was detected only in a few chronic
hepatitis patients. Clinical follow up study suggested that appearance of
anti-PreS1(21–47 aa) was related to the course of the disease and recovery of
patients. Detection system established in the study is promising for clinical
application.
Key words PreS1(21–47 aa) antigen peptide； GST-PreS1(21–47 aa) fusion
protein；
anti-PreS1(21–47 aa) antibody； biotin labeled protein A； ELISA
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