Construction of pMEKm12, an expression vector for protein production in Pseudomonas syringae

Abstract Characterization of the biological roles of proteins is essential for functional genomics of pseudomonads. Heterologous proteins overproduced in Escherichia coli frequently fail to exhibit biological function. To circumvent this problem, vector pMEKm12 was constructed and used to overexpres...

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Bibliographic Details
Published in:FEMS microbiology letters 2002-04, Vol.210 (1), p.115-121
Main Authors: Lu, Shi-En, Scholz-Schroeder, Brenda K., Gross, Dennis C.
Format: Article
Language:English
Subjects:
Bacterial Proteins - biosynthesis
Bacterial Proteins - genetics
Bacteriological methods and techniques used in bacteriology
Bacteriology
Biological and medical sciences
DNA, Bacterial - genetics
E coli
Escherichia coli - genetics
Fundamental and applied biological sciences. Psychology
Fusion protein
Genetic Vectors - biosynthesis
Genetics
Genomics
Maltose
Maltose-binding protein
Microbiology
Overexpression vector
Pathogens
Phytotoxin
Plasmids
Protein purification
Proteins
Pseudomonas
Pseudomonas - genetics
Pseudomonas syringae
Recombinant Fusion Proteins - biosynthesis
Replication Origin
Replication origins
Restoration
Restriction Mapping
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Summary:Abstract Characterization of the biological roles of proteins is essential for functional genomics of pseudomonads. Heterologous proteins overproduced in Escherichia coli frequently fail to exhibit biological function. To circumvent this problem, vector pMEKm12 was constructed and used to overexpress proteins in Pseudomonas. The vector contains the pRO1600 replication origin, the maltose-binding protein (MBP) fusion system, and an inducible tac promoter. The pMEKm12 was successfully used to overexpress the syringomycin synthetase SyrB1 protein fused to MBP in Pseudomonas syringae pv. syringae. Furthermore, expression of the MBP-SyrB1 protein in the syrB1 mutant BR132A1 resulted in the restoration of syringomycin production. This vector will facilitate confirmation of the biochemical roles of nonribosomal peptide synthetase genes in Pseudomonas syringae, and studies of gene function from a wide spectrum of pseudomonads.
ISSN:0378-1097
1574-6968
DOI:10.1111/j.1574-6968.2002.tb11169.x