Abstract
Aquaporin Z (AqpZ) is a typical orthodox aquaporin with 6 transmembrane domains and five connecting loops. In order to express this complex membrane protein efficiently, E. coli cell-free expression system was employed as an alternative to produce aquaporin Z. Using different fusion vectors containing AqpZ gene, the expression level of fusion proteins in cell-free system varied from 7.97 to 578.35 microg/ml, while 7.34 to 340.81 microg/ml for target protein (AqpZ). The free energy of mRNA secondary structure at translation initiation region (TIR) was predicted and demonstrated a positive relationship with the expression level of AqpZ in cell-free system. This is the first report of expressing water channel protein in E. coli cell-free system, which has become a highly promising tool for fast and efficient production of integral membrane proteins.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Aquaporins / biosynthesis*
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Aquaporins / chemistry
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Aquaporins / genetics*
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Binding Sites
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Biotechnology / methods
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Cell-Free System
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Enhancer Elements, Genetic
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Escherichia coli / genetics
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Escherichia coli / metabolism*
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Escherichia coli Proteins / biosynthesis*
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Escherichia coli Proteins / chemistry
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Escherichia coli Proteins / genetics*
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Escherichia coli Proteins / metabolism
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Gene Expression*
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Genetic Vectors*
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Histidine / genetics
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Histidine / metabolism
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Magnesium / chemistry
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Nucleic Acid Conformation
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Oligopeptides / genetics
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Oligopeptides / metabolism
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Osmolar Concentration
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RNA, Messenger / chemistry
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Recombinant Fusion Proteins / biosynthesis
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Recombinant Fusion Proteins / genetics
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Ribosomes
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Templates, Genetic
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Thioredoxins / genetics
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Thioredoxins / metabolism
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Transcription Initiation Site
Substances
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Aquaporins
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Escherichia coli Proteins
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His-His-His-His-His-His
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Oligopeptides
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RNA, Messenger
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Recombinant Fusion Proteins
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TrxA protein, E coli
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aqpZ protein, E coli
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Histidine
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Thioredoxins
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Magnesium