The 2-deoxystreptamine aglycon is a common structural feature found in aminocyclitol antibiotics including neomycm, kanamycin, tobramycin, gentamicin, sisomicin, butirosin and ribostamycin. A key enzyme involved in the biosynthesis of the 2-deoxystreptamine moiety is 2-deoxy-
scyllo-inosose (DOI) synthase which catalyses the carbocycle formation from D-glucose-6-phosphate to 2-deoxy-
scyllo-inosose. The recent success of isolating the 2-deoxy-
scyllo-inosose synthase from
Bacillus circulans prompted us to clone the gene responsible for this important enzyme by the use of reverse genetics approach. With the aid of DNA probes constructed on the basis of the amino-terminal sequence of the purified 42 kDa subunit of the enzyme, the responsible gene
btrC was successfully cloned. Subsequently the
btrC gene was heterologously expressed in
Escherichia coli, and the 2-deoxy-
scyllo-inosose synthase activity of the recombinant polypeptide was confirmed by chemical analysis. The
btrC gene encodes a protein composed of 368 amino acids with a molecular mass of 40.7 kDa. Our previous proposal for the similarity of 2-deoxy-
scyllo-inosose synthase to dehydroquinate synthase has been confirmed on the basis of their amino acid sequences. Significant differences in the sequences can also be observed however, particularly in the crucial substrate recognition regions. Comparison of the BtrC sequence with those of biosynthetic enzymes for other related microbial products is also discussed.
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