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Although the chemical structures of nikkomycins have been known since the 1970s, only a
few biosynthetic steps have been investigated in detail. The steps leading to the synthesis of
aminohexuronic acid are unclear. The biosynthesis commences with the attachment of an
enolpyruvyl moiety to the 3´-position of UMP. Since none of the nikkomycins synthesized
possess an enolpyruvyl group in this position of the sugar moiety, it must be concluded that
the resulting 3’-enolpyruvyl-UMP is subject to rearrangement reactions where the
enolpyruvyl is detached from its 3’-position and transferred to the 5’-position of the ensuing
aminohexuronic acid moiety. Co-crystallization of NikO with fosfomycin yielded rod – like
crystals diffracting up to 2.5 Å. A synchrotron dataset was measured at the Swiss Light
Source and the structure was solved by molecular replacement using UDP-Nacetylglucosamine
enolpyruvyl transferase (PDB code: 2rl1) as a model. Two molecules were
found in the asymmetric unit exhibiting an inverse α,β-barrel fold with helices forming the
tightly packed core and sheets shielding the hydrophobic core from the solvent:
Each chain is comprised of two inverse α,β-barrel subunits, which are connected by a hinge
region. The final structure was refined to final R/Rfree values of 17% and 19%, respectively.
The genes that are co-transcribed with nikO have been reported and are designated as
nikI, nikJ, nikK, nikL, nikM and nikN. In order to investigate the role of the encoded proteins
in the biosynthesis of the aminohexuronic acid moiety, these genes were cloned, expressed
and the proteins purified for biochemical characterization and crystallization. Furthermore,
nikkomycin intermediates produced by Streptomyces mutants featuring disruptions of the
respective genes were analyzed, but no nikkomycins could be detected in their fermentation
media. Comparisons of Nik enzyme sequences to those of known enzymes in the databases
allow some predictions of the reactions each nik enzyme might catalyze. None of the enzymes
was observed to convert 3´-EPUMP, the product of the NikO catalyzed reaction, so far. NikK
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