A novel geranylgeranyl reductase from the methanogenic archaeon Methanosarcina acetivorans displays unique regiospecificity.
Saturation of a prenyl group to various levels is a frequently observed modification of isoprenoids. The members of the geranylgeranyl reductase family, however, are the only known enzymes responsible for such reductive modifications in archaea. A methanogenic archaeon, Methanosarcina acetivorans, has proteins homologous to phytoene desaturase CrtI, which is the carotenogenic enzyme that catalyzes oxidation/isomerization of phytoene to lycopene, but their function in carotenogenesis is unlikely in a methanogen that does not produce carotenoids. In the present study, we identified one of the homologues, MA1492, as a new type of archaeal geranylgeranyl reductase that is not homologous to known geranylgeranyl reductases. The expression of MA1492 in Escherichia coli cells, which were genetically modified to produce unsaturated archaeal-type lipids, led to the production of partially saturated lipid derivatives. Furthermore, we analyzed the substrate specificity of recombinant MA1492 via in vitro assays. The LC-MS, or radio-TLC, analysis of the reaction products showed that the enzyme was definitely specific to compounds containing C20 geranylgeranyl groups and reduced only one of four double bonds in a geranylgeranyl chain. The GC-MS analysis of the product from geranylgeraniol confirmed that the reduction selectively occurred on the ω-terminal double bond. The available crystallographic structure of an orthologue enzyme may explain the reaction mechanism that achieves the substrate specificity and regiospecificity.
Department of Applied Molecular Bioscience, Graduate School of Bioagricultural Sciences, Nagoya University, Japan.
Takuya
T
Ogawa
Keisuke
K
Isobe
Takeshi
T
Mori
Susumu
S
Asakawa
Tohru
T
Yoshimura
Hisashi
H
Hemmi
D003001:Cloning, Molecular; D008401:Gas Chromatography-Mass Spectrometry; D017020:Methanosarcina / Q000201:enzymology*; D008958:Models, Molecular; D010088:Oxidoreductases / Q000737:chemistry* / Q000378:metabolism*; D011487:Protein Conformation; D013379:Substrate Specificity
D010088:Oxidoreductases; C110920:geranylgeranyl reductase
archaea; biosynthesis; geranylgeranyl reductase; isoprenoid; prenyl reductase