Surface display of HPV16L1 by autotransporter Ag43
10.3760/cma.j.cn112309-20211118-00391
- VernacularTitle:基于自转运蛋白Ag43的HPV16L1蛋白的细菌表面展示
- Author:
Kun CAI
1
;
Zhe WANG
;
Piying HUANG
;
Liangwan WEI
;
Xingyuan WANG
;
Xuemei XU
;
Yindi CHU
;
Peipei ZHU
;
Enguo FAN
Author Information
1. 中国医学科学院基础医学研究所北京协和医学院基础学院病原学系，医学分子生物学国家重点实验室，北京 100005
- Keywords:
Autotransporter;
Bacterial surface display;
HPV16L1;
Ag43
- From:
Chinese Journal of Microbiology and Immunology
2022;42(3):178-184
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To construct a surface display system containing various lengths of the Ag43 passenger domain for an optimal bacterial surface display of foreign protein HPV16L1.Methods:(1) Ag43 gene sequences of different lengths were inserted into pET22b vector to construct four Ag43 surface display vectors (Ag43/138, Ag43/551, Ag43/552 and Ag43/700) using PCR and subcloning strategy. (2) The generation of four HPV16L1-Ag43 fusion constructs was completed by PCR and subcloning methods. (3) HPV16L1-Ag43 fusion proteins were expressed and analyzed by SDS-PAGE. (4) The surface exposure of HPV-16L1 was verified using trypsin digestion.Results:PCR analysis and sequencing results showed that Ag43 surface display vectors and HPV16L1-Ag43 fusions were constructed successfully. SDS-PAGE showed that the expression of HPV16L1-Ag43 fusion proteins could be induced with 0.2 mmol/L IPTG and the protein content was reduced after the cells were treated with trypsin, especially the content of Ag43/700-HPV16L1 that showed a drastic reduction.Conclusions:The Ag43 surface display system was successfully constructed and could be used for a successful display of HPV16L1. This study also showed that Ag43/700 comprising only the α-helix and the β-barrel of Ag43 provided an optimal surface display for HPV16L1.
