Effects of Vitreoscilla hemoglobin on the 2,4-dinitrotoluene (2,4-DNT) dioxygenase activity of Burkholderia and on 2,4-DNT degradation in two-phase bioreactors

Expression of vgb, encoding Vitreoscilla hemoglobin (VHb), in Burkholderia strain YV1 was previously shown to improve cell growth and enhance 2,4-dinitrotoluene (2,4-DNT) degradation compared with control strain DNT, especially under hypoxic conditions. In the work reported here, the ratio of 2,4-DNT degraded to oxygen uptake was approximately 5-fold larger for strain YV1 than for strain DNT. The addition of purified VHb to cytosolic fractions of strain DNT increased 2,4-DNT degradation 1.5-fold, compared with 1.1-fold for control bovine Hb, but increased the 2,4-DNT degradation 2.7-fold when added to partially purified 2,4-DNT dioxygenase, compared with 1.3-fold for bovine Hb. This suggests a direct transfer of oxygen from VHb to the oxygenase. In a bioreactor at high 2,4-DNT concentration (using 100 ml oleyl alcohol containing 2 g 2,4-DNT as the second phase) with 1.5 l culture, both strains could remove 0.8 g 2,4-DNT by 120 h; and, under the same conditions in a fed-batch reactor, the degradation increased to 1 g for strain YV1 but not for strain DNT.