P64: Production of menaquinone-4 by Arthrobacter nicotianae

 Menaquinone, also known as vitamin K2, is involved in blood coagulation and bone metabolism and used as a dietary supplement in Japan. There exist a series of menaquinones having a various number of isoprene units in their side chain. Among them, menaquinone-4 (MK4) is known to be an active form of vitamin K2 in human since menaquinones with longer side chain are converted to MK4 in human body.

 Bacteria have menaquinones as an essential component of respiratory chain and produce menaquinones with longer side chain such as MK8. Arthrobacter nicotianae is known to produce MK8 and MK9 and a mutant strain of A. nicotianae producing mainly MK4 has been selected.

 Menaquinones are biosynthesized by the condensation of a quinone compound (1,4-dihydroxy-2-naphthoate, DHNA) and an isoprenoid compound such as geranylgeranyl pyrophosphate or octaprenyl pyrophosphate and the reaction is catalyzed by DHNA prenyltransferase (MenA). Genes involved in menaquinone biosynthesis were isolated from A. nicotianae and their effects on the length of menaquinone side chain were studied. When the gene of menA was overexpressed, the concentration of MK4 was specifically increased with no effects on that of menaquinones with longer side chain. When a plasmid expressing 1-deoxy-D-xylulose synthase (dxs) was introduced in order to increase a supply of isoprenoid, MK4 content was greatly reduced. These results indicated that the balance between MenA activity and the supply of isoprenoid was a key factor to control the length of menaquinone side chain.