Gene cloning, functional expression and secretion of the S-layer protein SgsE from Geobacillus stearothermophilus NRS 2004/3a in Lactococcus lactis
Gene cloning, functional expression and secretion of the S-layer protein SgsE from Geobacillus stearothermophilus NRS 2004/3a in Lactococcus lactis
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DOI：
10.1016/j.femsle.2004.10.036
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发表时间：
2005-01-01
影响因子：
2.1
通讯作者：
Schäffer, C
中科院分区：
文献类型：
--
作者：
Novotny, R;Scheberl, A;Schäffer, C
嗜热脂肪土芽孢杆菌NRS 2004/3a的类似于93-kDa的表面层蛋白SgsE形成规则的晶体阵列，为将来生物相关分子的展示提供纳米图案化的基质。乳酸乳球菌NZ 9000是一个安全的表达宿主，用于控制靶向生产SgsE的基础上广泛的宿主范围质粒pNZ 124 Sph，其中引入了nisA启动子。将缺乏其信号肽编码序列的SgsE克隆到新载体中，并以220 mg-1（-）表达培养物的产量从细胞质中纯化。分泌构建体基于短乳杆菌SlpA蛋白或L.乳酸菌Usp 45蛋白，允许分离95 mg分泌的rSgsE 1（-1）。N-末端测序证实了SgsE在L. lactis NZ9000.通过电子和原子力显微镜证实了rSgsE在悬浮液中自组装和在固体支持物上重结晶的能力。(C)2004年，欧洲微生物学会联合会。Elsevier B. V.出版，保留所有权利。
The similar to93-kDa surface layer protein SgsE of Geobacillus stearothermophilus NRS 2004/3a forms a regular crystalline array providing a nanopatterned matrix for the future display of biologically relevant molecules. Lactococcus lactis NZ9000 was established as a safe expression host for the controlled targeted production of SgsE based on the broad host-range plasmid pNZ124Sph, into which the nisA promoter was introduced. SgsE devoid of its signal peptide-encoding sequence was cloned into the new vector and purified from the cytoplasm at a yield of 220 mg 1(-) of expression culture. Secretion constructs were based on the signal peptide of the Lactobacillis brevis SlpA protein or the L. lactis Usp45 protein, allowing isolation of 95 mg of secreted rSgsE 1(-1). N-terminal sequencing confirmed correct processing of SgsE in L. lactis NZ9000. The ability of rSgsE to self-assemble in suspension and to recrystallize on solid supports was demonstrated by electron and atomic force microscopy. (C) 2004 Federation of European Microbiological Societies. Published by Elsevier B.V. All rights reserved.