Aspects related to mevalonate biosynthesis in plants

We purified and characterized a membrane‐associated enzyme system from radish (Raphanus sativus L.) that is capable of converting acetyl‐CoA into 3‐hydroxy‐3‐methylglutaryl‐coenzyme A (HMG‐CoA). The enzyme system apparently comprises acetoacetyl‐CoA thiolase (EC 2.3.1.9) and HMG‐CoA synthase (EC 4.1.3.5). Its activityin vitro can be strongly stimulated by FeII. When ferrous ions are applied chelated with ethylenediaminetetraacetate, citrate or adenosine 5′‐triphosphate (ATP), the stimulation is further increased. Stimulation is due to a higher catalytic efficiency as indicated by an increase in Vmax, whereas the affinity of the enzyme towards acetyl‐CoA remains constant (Km=6 μM). A considerable portion of HMG‐CoA lyase activity is associated with the same membranes. HMG‐CoA lyase (EC 4.1.3.4) is also solubilized and partially co‐purified. Its activity requires comparatively high concentrations of Mg2+. The conversion of HMG‐CoA to mevalonic acid is catalyzed by HMG‐CoA reductase (EC 1.1.1.34) that is associated with the same membranes. By cDNA encoding theArabidopsis HMG‐CoA reductase, we isolated a corresponding gene from a cDNA library newly established from etiolated radish seedlings. This full‐length cDNA, referred to as λcRS3, encodes a polypeptide of 583 amino acids with a molecular mass of about 63 kDa. The hydropathy profile suggests the presence of two hydrophobic membrane‐spanning domains within the N‐terminal 165 amino acids. The carboxy‐terminal part, where the catalytic site resides, is highly conserved in all eukaryotic HMG‐CoA reductase genes sequenced so far.