The enhancement of microorganisms for chemical modification

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Aromatic hydrocarbon utilizing bacteria have become an increased concern in the aspect of environmental bioremediation and production of specific chemicals. The present research has been conducted in a collaboration with University College London. The aim of this research is to enhance microorganisms for degradation and modification of specific chemicals. The results obtained in the joint research are as follows.1. The structure and function of bphABCXD genes of Pseudomonas pseudoalcaligenes KF707 coding for conversion of biphenyl to benzoic acid were analyzed. Biphenyl is oxidized to dihydrodio by a multi-component enzyme, biphenyl dioxygenase which is composed of four different subunits : large and small subunits of terminal dioxygenase (encoded by bphA1 and bphA2), ferredoxin (bphA3) and ferredoxin reductase (bphA4). The dihydrodiol is then dihydrogenated to 2,3-dihydroxybiphneyl by a dehydrogenase (bphB). 2,3-dihydroxybiphenyl dioxgenase (bphc) convert 2,3-dihydroxybiphenyl into th … More e ring-meta-cleavage compound, 2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoate, which is eventually degraded to benzoic acid by a hydrolase (bphD).2. A toluene catabolic tod operon of P. putida F1 was analyzed by Gibson's group. It was found that bph and tod operon are very similar, not only in the gene organization, but also the size and amino acid sequences of the corresponding gene components. Thus' identities of amino acid sequences between bphA1/todC1, bphA2/todC2, bphA3/todB, and bphC/todE are 53-65%. Biphenyl utilizing KF707 is not able to utilize toluene. It was demonstrated that introduction of plasmid pDTG351 containing todC1C2BADE genes into the KF707 allowed the recombinant strain to grow on toluene. More importantly, introduction of only todC1C2 supported the growth of the KF707 on toluene.3. Using transposon mutant strains, KF748 (bphB :: Tn5-B21) and KF744 (bphC :: Tn5-B21), dihydrodiol and dihydroxy compounds were produced from various biphenyl derivatives and biphenyl-related compounds, indicating bph-encoded enzymes possess very relaxed substrate specificities. Less