Cloning, expression, and characterization of the Anabaena thioredoxin gene in Escherichia coli
Cloning, expression, and characterization of the Anabaena thioredoxin gene in Escherichia coli
复制标题
大肠杆菌中鱼腥藻硫氧还蛋白基因的克隆、表达和表征
作者：
C. Lim;F. K. Gleason;J. Fuchs
基于两种硫氧还蛋白之间的相似性将反映在基因序列和功能交叉反应性上的策略，在大肠杆菌中克隆鱼腥藻菌株PCC 7119中的硫氧还蛋白编码基因。通过与大肠杆菌异源杂交，鉴定了含有鱼腥藻硫氧还蛋白基因的DNA限制性片段。coli硫氧还蛋白基因，与pUC 13连接，转化大肠杆菌。缺乏功能性硫氧还蛋白的大肠杆菌菌株。在E.大肠杆菌通过增加的菌落大小鉴定，并通过酶测定确认。鱼腥藻硫氧还蛋白基因在大肠杆菌中的表达。大肠杆菌中的藻蛋白的纯化和鉴定证实了这一点。杆菌来源于鱼腥藻基因DNA序列的氨基酸序列与鱼腥藻硫氧还蛋白的已知氨基酸序列相同。急诊表达鱼腥藻硫氧还蛋白的大肠杆菌菌株在各方面都补充了TrxA-表型，除了它们不支持噬菌体T7生长并且支持噬菌体M13和f1的能力有所降低。
The gene encoding thioredoxin in Anabaena sp. strain PCC 7119 was cloned in Escherichia coli based on the strategy that similarity between the two thioredoxins would be reflected both in the gene sequence and in functional cross-reactivity. DNA restriction fragments containing the Anabaena thioredoxin gene were identified by heterologous hybridization to the E. coli thioredoxin gene following Southern transfer, ligated with pUC13, and used to transform an E. coli strain lacking functional thioredoxin. Transformants that complemented the trxA mutation in E. coli were identified by increased colony size and confirmed by enzyme assay. Expression of the cloned Anabaena thioredoxin gene in E. coli was substantiated by subsequent purification and characterization of the algal protein from E. coli. The amino acid sequence derived from the DNA sequence of the Anabaena gene was identical to the known amino acid sequence of Anabaena thioredoxin. The E. coli strains which expressed Anabaena thioredoxin complemented the TrxA- phenotype in every respect except that they did not support bacteriophage T7 growth and had somewhat decreased ability to support bacteriophages M13 and f1.
蓝藻鱼腥藻中硫氧还蛋白的分离和表征。
DOI：
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发表时间：
1981
期刊：
The Journal of biological chemistry
影响因子：
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作者：
Gleason,FK;Holmgren,A
通讯作者：
Holmgren,A
DOI：
10.1073/pnas.78.6.3393
发表时间：
1981-01-01
期刊：
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA-BIOLOGICAL SCIENCES
影响因子：
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作者：
FISHER, R;TULI, R;HASELKORN, R
通讯作者：
HASELKORN, R