Abstract
The inhibition of mevalonate kinase (MVK) by downstream metabolites is an important mechanism in the regulation of isoprenoid production in a broad range of organisms. The first feedback-resistant MVK was previously discovered in the methanogenic archaeon Methanosarcinamazei. Here, we report the cloning, expression, purification, kinetic characterization and inhibition analysis of MVKs from two other methanogens, Methanosaetaconcilii and Methanocellapaludicola. Similar to the M. mazei MVK, these enzymes were not inhibited by diphosphomevalonate (DPM), dimethylallyl diphosphate (DMAPP), isopentenyldiphosphate (IPP), geranylpyrophosphate (GPP) or farnesylpyrophosphate (FPP). However, they exhibited significantly higher affinity to mevalonate and higher catalytic efficiency than the previously characterized enzyme.
Highlights
Isoprenoids, or terpenoids, are a very large class of naturally occurring compounds with diverse structures and functions
Since putative genes encoding phosphomevalonate kinase (PMK) and diphosphomevalonate decarboxylase (DMD) were not identified on the basis of sequence similarity in archaeal genomes, while isopentenyl phosphate kinase (IPK) genes have been found in most archaea, it has been hypothesized that an alternative mevalonate pathway operates in Archaea [12]
We report here the cloning and overexpression of the mvk genes from Methanosaeta concilii, an archaeon belonging to the same order, Methanosarcinales, as M. mazei, from an evolutionarily distant methanogenic archaeon, Methanocella paludicola, and from Nitrosopumilus maritimus, an unrelated species belonging to Thaumarchaeota, in Escherichia coli; the purification, kinetic characterization and inhibition analysis of the corresponding enzymes are described
Summary
Isoprenoids, or terpenoids, are a very large class of naturally occurring compounds with diverse structures and functions. Since putative genes encoding phosphomevalonate kinase (PMK) and diphosphomevalonate decarboxylase (DMD) were not identified on the basis of sequence similarity in archaeal genomes, while isopentenyl phosphate kinase (IPK) genes have been found in most archaea, it has been hypothesized that an alternative mevalonate pathway operates in Archaea [12]. In this hypothetical pathway, phosphomevalonate is decarboxylated to yield isopentenyl phosphate (IP), which is phosphorylated to IPP by the action of IPK. MVA-3-P, but not MVA-5-P, serves as a precursor for IP synthesis in Thermoplasma acidophilum; a novel enzyme, ATP:(R)-MVA 3-phosphotransferase Ta1305, was identified in this species [16]
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