UDP-glucose:: (6-methoxy)podophyllotoxin 7-O-glucosyltransferase from suspension cultures of Linum nodiflorum

Cell cultures of Linum species store 6-methoxypodophyllotoxin (MPTOX), podophyllotoxin (PTOX) and related lignans as O-glucosides. UDP-glucose:(M)PTOX 7-O-glucosyltransferase has been detected and characterised in protein preparations of suspension-cultured cells of Linum nodiflorum L. (Linaceac). The maximal lignan glucoside contents in the cells are preceded by a rapid increase of the specific glucosyltransferase, activity on day six of the culture period. MPTOX glucoside is the major lignan with up to 1.18 mg g(-1) of the cell dry wt which is more than 30-fold of the PTOX glucoside content. Of the three aryltetralin lignans tested as substrates, PTOX and MPTOX display comparable apparent K-m values of 4.7 and 5.4 mu M, respectively. 5'-Demethoxy-6-methoxypodophyllotoxin is converted with the highest velocity of 25.2 pkat mg(-1) while also possessing a higher K-m of 14.7 mu M. Two-substrate test series indicate that all three compounds compete for the active site of a single protein. The structurally similar lignan O-peltatin acts as competitive inhibitor as well. However, the 6-O-glucosidation is most likely catalysed by a separate enzyme. The (M)PTOX 7-O-glucosyltransferase works best at a pH around 9 and a temperature around 35 degrees C. A 15-30% increase of the reaction rate is effected by the addition of 0.9 mM Mn2+. (C) 2007 Elsevier Ltd. All rights reserved.