Expression of a cloned beta-glucanase gene from Bacillus amyloliquefaciens in an Escherichia coli relA strain after plasmid amplification.

TY - JOUR

T1 - Expression of a cloned beta-glucanase gene from Bacillus amyloliquefaciens in an Escherichia coli relA strain after plasmid amplification.

AU - Hecker, M

AU - Riethdorf, Sabine

AU - Bauer, C

AU - Schroeter, A

AU - Borriss, R

PY - 1988

Y1 - 1988

N2 - Amino acid starvation of cells of the Escherichia coli relA strain, CP79, which cannot accumulate guanosine tetraphosphate (ppGpp) in response to amino acid limitation, increased the pEG1 plasmid content about 5- to 7-fold in comparison with exponentially growing cells (pEG1:pBR322 with an insertion of Bacillus amyloliquefaciens DNA coding for beta-glucanase). In contrast, no pEG1 amplification occurred in E. coli CP78, the stringently controlled counterpart, after amino acid starvation. In order to verify these results, the plasmid DNA content was monitored by measuring the expression of pEG1-encoded beta-glucanase from B. amyloliquefaciens both before and after plasmid amplification. When amino acid starved CP79 cells were given an additional dose of amino acids, a more than 10-fold increase in pEG1-encoded beta-glucanase activity (per cell mass) was measured. This increase in enzyme activity correlates with pEG1 amplification during amino acid limitation. Under comparable conditions the activity of beta-glucanase was not increased in strain CP78, which did not amplify the plasmid. We suggest that the replication of pEG1 in amino acid starved E. coli cells is somehow under negative control by ppGpp. Moreover, we found the Bacillus beta-glucanase in E. coli relA cells to be excreted into the growth medium after starvation and overexpression.

AB - Amino acid starvation of cells of the Escherichia coli relA strain, CP79, which cannot accumulate guanosine tetraphosphate (ppGpp) in response to amino acid limitation, increased the pEG1 plasmid content about 5- to 7-fold in comparison with exponentially growing cells (pEG1:pBR322 with an insertion of Bacillus amyloliquefaciens DNA coding for beta-glucanase). In contrast, no pEG1 amplification occurred in E. coli CP78, the stringently controlled counterpart, after amino acid starvation. In order to verify these results, the plasmid DNA content was monitored by measuring the expression of pEG1-encoded beta-glucanase from B. amyloliquefaciens both before and after plasmid amplification. When amino acid starved CP79 cells were given an additional dose of amino acids, a more than 10-fold increase in pEG1-encoded beta-glucanase activity (per cell mass) was measured. This increase in enzyme activity correlates with pEG1 amplification during amino acid limitation. Under comparable conditions the activity of beta-glucanase was not increased in strain CP78, which did not amplify the plasmid. We suggest that the replication of pEG1 in amino acid starved E. coli cells is somehow under negative control by ppGpp. Moreover, we found the Bacillus beta-glucanase in E. coli relA cells to be excreted into the growth medium after starvation and overexpression.

KW - Gene Expression Regulation

KW - Gene Amplification

KW - Cloning, Molecular

KW - Plasmids

KW - Amino Acids/pharmacology

KW - Genes, Bacterial

KW - Bacillus/enzymology/genetics

KW - Escherichia coli/drug effects/enzymology/genetics

KW - Glycoside Hydrolases/genetics

KW - Guanosine Tetraphosphate/genetics

KW - Gene Expression Regulation

KW - Gene Amplification

KW - Cloning, Molecular

KW - Plasmids

KW - Amino Acids/pharmacology

KW - Genes, Bacterial

KW - Bacillus/enzymology/genetics

KW - Escherichia coli/drug effects/enzymology/genetics

KW - Glycoside Hydrolases/genetics

KW - Guanosine Tetraphosphate/genetics

M3 - SCORING: Journal article

VL - 215

SP - 181

EP - 183

JO - Mol Gen Genet

JF - Mol Gen Genet

SN - 0026-8925

IS - 1

M1 - 1

ER -