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A metal‐repressed promoter from gram‐positive Bacillus subtilis is highly active and metal‐induced in gram‐negative Cupriavidus metallidurans
Author(s) -
RibeirodosSantos Gabriela,
Biondo Ronaldo,
Quadros Oeber de Freitas,
Vicente Elisabete José,
Schenberg Ana Clara Guerrini
Publication year - 2010
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/bit.22820
Subject(s) - bacillus subtilis , escherichia coli , heterologous , promoter , bacteria , microbiology and biotechnology , gene , chemistry , reporter gene , gene expression , biology , zinc , biochemistry , genetics , organic chemistry
A synthetic version of the metal‐regulated gene A ( mrgA ) promoter from Bacillus subtilis , which in this Gram‐positive bacterium is negatively regulated by manganese, iron, cobalt, or copper turned out to promote high level of basal gene expression that is further enhanced by Co(II), Cd(II), Mn(II), Zn(II), Cu(II), or Ni(II), when cloned in the Gram‐negative bacterium Cupriavidus metallidurans . Promoter activity was monitored by expression of the reporter gene coding for the enhanced green fluorescent protein (EGFP), and cellular intensity fluorescence was quantified by flow cytometry. Expression levels in C. metallidurans driven by the heterologous promoter, here called pan , ranged from 20‐ to 53‐fold the expression level driven by the Escherichia coli lac promoter (which is constitutively expressed in C. metallidurans ), whether in the absence or presence of metal ions, respectively. The pan promoter did also function in E. coli in a constitutive pattern, regardless of the presence of Mn(II) or Fe(II). In conclusion, the pan promoter proved to be a powerful tool to express heterologous proteins in Gram‐negative bacteria, especially in C. metallidurans grown upon high levels of toxic metals, with potential applications in bioremediation. Biotechnol. Bioeng. 2010;107: 469–477. © 2010 Wiley Periodicals, Inc.