Wei sheng wu xue bao = Acta microbiologica Sinica · 2008
[Incorporation of phosphatidylcholine into Escherichia coli membrane affects secretion of penicillin beta-lactamase].
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Cai Xueli, Li Yang, Xuan Wenjing, Wang Xinguo
Аннотация
OBJECTIVE: To study the biological function of phosphatidylcholine in bacteria, the borrelial pcs gene was inserted into ptac85 plasmid. Then E. coli Top10 pcs+ was constructed via the transformation of the recombinant plasmid. Phosphatidylcholine (30%) in total phospholipids was achieved when the bacterial cells were incubated in Luria-Bertani (LB) medium supplemented with 1% choline and induced by 0.5 mmol/L isopropy-beta-D-thiogalactoside (IPTG) for 4-8 hours at 37 degrees C.
METHODS: Ampicillin inhibitionof E. coli Top10 pcs+ was tested at first, and then beta-lactamase activity in periplasm was examined. Finally Western blot was used to detect the amount of beta-lactamase in both bacterial periplasm and cytoplasm.
RESULTS: Antibiotic tests showed that high concentrations of ampicillin inhibited the growth of E. coli Top100 pcs+ with an IC50 of 70-800 microg/mL. Active assays revealed that the beta-lactamase activity in periplasm was only 1/5 of that for the control strain E. coli Top10/p(tac)85. Western blotting confirmed that the low activity of beta-lactamase in E. coli Top10 pcs+ resulted from a lower amount of beta-lactamase in its periplasm.
CONCLUSION: Our results demonstrated that the phospatidylcholine incorporated into bacterial membrane retarded secretion of Escherichia coli penicillin beta-lactamase from cytoplasm into periplasm, which suggested that phosphatidylcholine might play a role in the regulation of protein secretion in those bacteria able to synthesize phosphatidylcholine.
METHODS: Ampicillin inhibitionof E. coli Top10 pcs+ was tested at first, and then beta-lactamase activity in periplasm was examined. Finally Western blot was used to detect the amount of beta-lactamase in both bacterial periplasm and cytoplasm.
RESULTS: Antibiotic tests showed that high concentrations of ampicillin inhibited the growth of E. coli Top100 pcs+ with an IC50 of 70-800 microg/mL. Active assays revealed that the beta-lactamase activity in periplasm was only 1/5 of that for the control strain E. coli Top10/p(tac)85. Western blotting confirmed that the low activity of beta-lactamase in E. coli Top10 pcs+ resulted from a lower amount of beta-lactamase in its periplasm.
CONCLUSION: Our results demonstrated that the phospatidylcholine incorporated into bacterial membrane retarded secretion of Escherichia coli penicillin beta-lactamase from cytoplasm into periplasm, which suggested that phosphatidylcholine might play a role in the regulation of protein secretion in those bacteria able to synthesize phosphatidylcholine.