A gene cluster encoding five enzymes of the mevalonate pathway had
been cloned from
Streptomyces
sp. strain CL190. This
gene cluster contained an additional ORF,
orfD
, encoding
an unknown protein that was detected in some archaebacteria and some
Gram-positive bacteria including
Staphylococcus aureus
.
The recombinant product of
orfD
was purified as a
soluble protein and characterized. The molecular mass of the enzyme was
estimated to be 37 kDa by SDS-polyacrylamide gel electrophoresis and
155 kDa by gel filtration chromatography, suggesting that the enzyme is
most likely to be a tetramer. The purified enzyme contained flavin
mononucleotide (FMN) with the amount per tetramer being 1.4 to 1.6
mol/mol. The enzyme catalyzed the isomerization of isopentenyl
diphosphate (IPP) to produce dimethylallyl diphosphate (DMAPP) in the
presence of both FMN and NADPH. The
Escherichia coli
plasmid expressing
orfD
could complement the disrupted
IPP isomerase gene in
E. coli
. These results indicate
that
orfD
encodes an unusual IPP isomerase showing no
sequence similarity to those of IPP isomerases identified to date.
Based on the difference in enzymatic properties, we classify the IPP
isomerases into two types: Type 2 for FMN- and NAD(P)H-dependent
enzymes, and type 1 for the others. In view of the critical role of
this isomerase in
S. aureus
and of the different
enzymatic properties of mammalian (type 1) and
S. aureus
(type 2) isomerases, this unusual enzyme is considered to be a suitable
molecular target for the screening of antibacterial drugs specific to
S. aureus
.
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