Microbial Demetallization of Crude Oil : Nickel Protoporphyrin Disodium as a Model Organo-Metallic Substrate

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A soil isolate designated as YA-1 strain was selected for its ability to degrade nickel protoporphyrin disodium(NiPPDS). The strain was capable of utilizing NiPPDS as the sole source of carbon. This strain, a gram-negative aerobic rod, was identified as Pseudomonas azelaica YA-1 based on the result of its 16S rRNA analysis. Product analyses by HPLC showed that this strain can decompose the porphyrin ring to which a metal ion is bound. However, the use of whole bacterial cells cannot result in extensive NiPPDS degradation; therefore, the YA-1 enzyme was extracted and purified. This NiPPDS-degrading enzyme named as protoporphyrinase was purified from P.azelaica YA-1 by ammonium sulfate fractionation and sequential chromatographies using DEAE Toyopearl 650 M, CM Toyopearl 650 M and Biogel P-60 columns, with a yield of 11.3% based on the enzyme activity and an overall purification of 498-fold. The molecular weight of this enzyme is estimated to be 39, 000 Da by SDS-PAGE and 34, 000 Da by gel filtration. The optimum pH and temperature for the enzyme were 7.0 and 30℃, respectively. The activity was stable at pH 2.0-11.0 and at temperatures below 50℃. The enzyme activity was inactivated by ferric chloride, potassium ferricyanide, ZnCl_2 and CdCl_2.
社団法人日本生物工学会の論文
2000-11-25