EXPRESSION OF A CLONED CELLULASE XYLANASE GENE FROM PREVOTELLA-RUMINICOLA IN BACTEROIDES-VULGATUS, BACTEROIDES UNIFORMIS AND PREVOTELLA-RUMINICOLA
EXPRESSION OF A CLONED CELLULASE XYLANASE GENE FROM PREVOTELLA-RUMINICOLA IN BACTEROIDES-VULGATUS, BACTEROIDES UNIFORMIS AND PREVOTELLA-RUMINICOLA
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DOI：
10.1111/j.1365-2672.1995.tb03156.x
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发表时间：
1995-10-01
期刊：
影响因子：
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通讯作者：
FLINT, HJ
中科院分区：
文献类型：
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作者：
DANIEL, AS;MARTIN, J;FLINT, HJ
构建了一个新的穿梭载体pRH3(8.7kb)，用于普氏杆菌/类杆菌宿主菌株。该载体将瘤胃假单胞菌的pRR12复制子、pBluescrip序列和一个ETQ标记基因相结合，用于普氏杆菌/类杆菌宿主的筛选。将来自瘤胃假单胞菌23的携带内切葡聚糖酶/木聚糖酶基因的片段插入多克隆位点后，得到的构建物pRH3X被导入到普通芽孢杆菌1447、均匀双胞菌1100和瘤胃假单胞菌2202中。这导致在葡萄糖生长的细胞中CM-纤维素酶和木聚糖酶的活性增加了4到50倍。相比之下，在E.ColiDH5α中几乎检测不到相同结构的活性。PRH3X转化瘤胃拟青霉2202和普通芽孢杆菌1447产生的木聚糖酶主要在细胞内产生，而在瘤胃拟青霉23中，渗透压休克实验表明，携带pRH3X的普通芽孢杆菌1447细胞的木聚糖酶活力有很大比例是周质的。
A new shuttle vector, pRH3 (8.7 kb), was constructed for use in Prevotella/Bacteroides host strains. This vector combines the pRR12 replicon from P. ruminicola, pBluescript sequences and a tetQ marker gene for selection in Prevotella/Bacteroides hosts. Following insertion of a fragment carrying an endoglucanase/xylanase gene from P. ruminicola 23 into the multiple cloning site, the resulting construct, pRH3X, was introduced into B. vulgatus 1447, B. uniformis 1100 and P. ruminicola 2202. This resulted in increases of between 4 and 50-fold in CM-cellulase and xylanase activities in cells grown with glucose. In contrast activities were barely detectable for the same construct in E. coli DH5 alpha. Most of the total xylanase activity produced was found within the cell in P. ruminicola 2202 and B. vulgatus 1447 transformed with pRH3X, and in P. ruminicola 23, An osmotic shock experiment indicated that a significant proportion of the xylanase activity in B. vulgatus 1447 cells carrying pRH3X was periplasmic.