Double mutation of cell wall proteins CspB and PBP1a increases secretion of the antibody Fab fragment from Corynebacterium glutamicum.
Double mutation of cell wall proteins CspB and PBP1a increases secretion of the antibody Fab fragment from Corynebacterium glutamicum.
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细胞壁蛋白 CspB 和 PBP1a 的双重突变增加了谷氨酸棒杆菌抗体 Fab 片段的分泌。
DOI：
10.1186/1475-2859-13-56
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发表时间：
2014-04-15
影响因子：
6.4
通讯作者：
Wachi M
中科院分区：
文献类型：
--
作者：
Matsuda Y;Itaya H;Kitahara Y;Theresia NM;Kutukova EA;Yomantas YA;Date M;Kikuchi Y;Wachi M
Among other advantages, recombinant antibody-binding fragments (Fabs) hold great clinical and commercial potential, owing to their efficient tissue penetration compared to that of full-length IgGs. Although production of recombinant Fab using microbial expression systems has been reported, yields of active Fab have not been satisfactory. We recently developed the Corynebacterium glutamicum protein expression system (CORYNEX®) and demonstrated improved yield and purity for some applications, although the system has not been applied to Fab production. The Fab fragment of human anti-HER2 was successfully secreted by the CORYNEX® system using the conventional C. glutamicum strain YDK010, but the productivity was very low. To improve the secretion efficiency, we investigated the effects of deleting cell wall-related genes. Fab secretion was increased 5.2 times by deletion of pbp1a, encoding one of the penicillin-binding proteins (PBP1a), mediating cell wall peptidoglycan (PG) synthesis. However, this Δpbp1a mutation did not improve Fab secretion in the wild-type ATCC13869 strain. Because YDK010 carries a mutation in the cspB gene encoding a surface (S)-layer protein, we evaluated the effect of ΔcspB mutation on Fab secretion from ATCC13869. The Δpbp1a mutation showed a positive effect on Fab secretion only in combination with the ΔcspB mutation. The ΔcspBΔpbp1a double mutant showed much greater sensitivity to lysozyme than either single mutant or the wild-type strain, suggesting that these mutations reduced cell wall resistance to protein secretion. There are at least two crucial permeability barriers to Fab secretion in the cell surface structure of C. glutamicum, the PG layer, and the S-layer. The ΔcspBΔpbp1a double mutant allows efficient Fab production using the CORYNEX® system.
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影响因子：
2.7
作者：
Burtet, Rafael Trindade;Santos-Silva, Marcos Antonio;Brigido, Marcelo Macedo
通讯作者：
Brigido, Marcelo Macedo
影响因子：
3.2
作者：
JARLIER, V;NIKAIDO, H
通讯作者：
NIKAIDO, H
影响因子：
6.4
作者：
BLASER, MJ;SMITH, PF;WANG, WLL
通讯作者：
WANG, WLL
影响因子：
4.1
作者：
Hermann, T
通讯作者：
Hermann, T
影响因子：
4.4
作者：
Kikuchi, Y;Date, M;Matsui, H
通讯作者：
Matsui, H