An unusual isopentenyl diphosphate isomerase found in the mevalonate pathway gene cluster from Streptomyces sp. strain CL190

Abstract

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 .

Identification of a fourth family of lycopene cyclases in photosynthetic bacteria

“…It is possible that its catalytic function is most similar to that of the type II isopentenyl diphosphate (IPP)/ dimethylallyl diphosphate (DMAPP) isomerase, which converts IPP to DMAPP by isomerizing a double bond (62,63). Both type I and type II IPP/DMAPP isomerases appear to catalyze isomerization by stereoselective protonation and deprotonation of the substrate (64,65), and the type II IPP isomerase, like CrtY, requires a reduced flavin and NADPH for activity (62,63,64). In this case, although there is no net oxidation or reduction, both reduction and oxidation of the substrate occur during the reaction, and the flavin may have a role in these half-reactions.…”

New Role of Flavin as a General Acid-Base Catalyst with No Redox Function in Type 2 Isopentenyl-diphosphate Isomerase

“…Type 2 isopentenyl-diphosphate isomerase (IDI) 4 is the flavoenzyme that catalyzes the interconversion between isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP) that occurs with no net change in redox status (5). Both compounds are fundamental units for the biosynthesis of isoprenoids, a diverse family of Ͼ50,000 metabolites (6).…”

Efficient Production of δ-Guaiene, an Aroma Sesquiterpene Compound Accumulated in Agarwood, by Mevalonate Pathway-Engineered <i>Escherichia coli</i> Cells

“…Recently, Harada et al [12] constructed MVA pathway-engineered E. coli cells by the transformation with a gene cluster ( Figure 2) isolated from Streptomyces sp. [13] [14], and demonstrated [15] that efficient carotenoid production was achieved in this strain by overexpression of several genes involved in the biosynthetic processes of the tetraterpene pigments. In the present experiments, we have attempted to construct the efficient producing system of δ-guaiene, which is biosynthesized in Aquilaria plants only under very limited condition, employing MVA pathway-engineered E. coli cells.…”