Dynamic interaction between components of hexaprenyl diphosphate synthase from Micrococcus luteus BP-26.
Hexaprenyl diphosphate synthase from Micrococcus luteus BP-26, which has been known to be dissociated into two essential components, designated as components A and B, during hydroxyapatite chromatography [Fujii, H., Koyama, T., & Ogura, K. (1982) J. Biol. Chem. 257, 14610-14612], was also resolved similarly by Sephadex G-100 or DEAE ion-exchange chromatography. Each component takes various self-aggregated forms. The apparent molecular mass of component B estimated by gel filtration on Superose 12 varied depending on its concentration, ranging from approximately 18 to 49 kilodaltons (kDa). On the other hand, the apparent molecular mass of component A varied depending on not only its concentration but also the ionic strength of the medium, ranging from approximately 13 to 24 kDa. When a mixture of components A and B preincubated in the presence of Mg2+ but in the absence of substrate was subjected to Superose 12 gel filtration, they were eluted at positions identical with those observed when they were chromatographed individually. In contrast, when a mixture of components A and B incubated in the presence of Mg2+ and substrates was filtrated on Superose 12, the elution positions were markedly changed, showing that an approximately 24-kDa aggregate of component A (designated as A) and an approximately 27-kDa aggregate of component B (designated as B) were the major species. Evidence was also obtained to show that farnesyl diphosphate (FPP) binds to the components to form an aggregate, A-B-FPP-Mg2+, which probably represents an intermediary state of enzyme catalysis.
Yoshida
I
I
Chemical Research Institute of Non-Aqueous Solutions, Tohoku University, Sendai, Japan.
Koyama
T
T
Ogura
K
K
D019883:Alkyl and Aryl Transferases; D002845:Chromatography; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004122:Dimethylallyltranstransferase / Q000302:isolation & purification / Q000378:metabolism*; D017886:Durapatite; D006882:Hydroxyapatites; D007700:Kinetics; D046911:Macromolecular Substances; D008837:Micrococcus / Q000201:enzymology*; D008970:Molecular Weight; D014166:Transferases / Q000378:metabolism*
D006882:Hydroxyapatites; D046911:Macromolecular Substances; D017886:Durapatite; D014166:Transferases; D019883:Alkyl and Aryl Transferases; D004122:Dimethylallyltranstransferase; C036826:trans-pentaprenyltranstransferase