Effect of Vitreoscilla hemoglobin on production anti-tumor enzyme methionine –γ-lyase, by Citrobacter freundii
Methionine, a sulfur amino acid, is the first amino acid that is required for many proteins, during synthesis. Our preliminary studies showed that this compound was produced during the late (post-stationary) secondary phase of growth. Therefore, restriction of methionine may be a useful strategy in limiting cancer growth. The bacterial strain used in this study was Citrobacter freundii (NRRL B-2643) and their vgb+ recombinant strain. A 1/100 inoculum of overnight cultures grown in LB was made in 50 ml LB in 150 ml Erlenmeyer flasks. Inocula in flasks were grown for 24 h at 30 °C in a 200 rpm water-bath. For MGL production, 250 μL of this O/N culture was then inoculated into 150 mL conical flask containing 50 mL of sterile mineral salts medium supplemented with 1 % or 0.1 % (w/v) glucose, respectively. This was incubated for 96 h at 30 °C, 200 rpm on an orbital shaker. The highest MGL concentration (2,02) was reached by the recombinant strain of Cf[pUC8:15] 72 h after the start of incubation MM+0,1% glucose source. In comparison, the wild type strain produced 3,14 of MGL concentration 72 h was reached MM+0,1% glucose source. The poor media and secondary phase (72 h and up) was used to for MGL production. This is more appropriate. Plasmid is disadvantages in the secondary stage.
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