Stereoselective biosynthesis of chloroarylpropane diols by the basidiomycete Bjerkandera adusta: exploring the roles of amino acids, pyruvate, glycerol and phenyl acetyl carbinol

Abstract

Bjerkandera adusta produces many chlorometabolites including chlorinated anisyl metabolites (CAMs) and 1-arylpropane-1,2-diols ( 1, 2, 3, 4) as idiophasic metabolic products of L-phenylalanine. These diols are stereoselectively biosynthesized from a C 7-unit (benzylic, from L-phenylalanine) and a C 2-unit, of unknown origin, as predominantly erythro (1 R,2 S) enantiomers. Of the labeled amino acids tested as possible C 2-units, at the 4–10 mM level, none were found to efficiently label the 2,3-propane carbons of the diols. However, glycine (2- 13C), L-serine (2,3,3-d 3) and L-methionine (methyl-d 3) entered the biomethylation pathway. Neither pyruvate (2,3- 13C 2), acetate (1,2- 13C 2), acetaldehyde (d 4) nor ethanol (ethyl-d 5) labeled the 2,3-propane carbons of the diols at the 4–10 mM level. Pyruvate (2,3- 13C 2) and L-serine (2,3,3-d 3) (which also entered the biomethylation pathway) did, however, effectively label the 2,3-propane carbons of the α-ketols and diols at the 40 mM level as evidenced by mass spectrometry. Glycerol (1,1,2,3,3-d 5) also appeared to label one of the 2,3-propane carbons (ca. 5% as 2H 2 in the C3 side chain) as suggested by mass spectrometric data and also entered the biomethylation pathway, likely via amino acid synthesis. Glycerol (through pyruvate), therefore, likely supplies C2 and C3 of the propane side chain with arylpropane diol biosynthesis. Incubation of B. adusta with synthetic [2- 2H 1,2- 18O]-glycerol showed that neither 2H nor 18O were incorporated in the α-ketols or diols. The oxygen atom on the C2 of the ketols/diols, therefore, does not appear to come from the oxygen atom on the C2 of glycerol. Glycerol, however, can readily form L-serine (which can then form pyruvate via PLP/serine dehydratase and involve transamination washing out the 18O label and providing the oxygen from water), and can then go on to label the C 2-unit. Labeled α-ketol, phenyl acetyl carbinol ( 5) (PAC; ring-d 5, 2,3- 13C 2 propane) cultured with B. adusta leads to stereospecific reduction to the (1 R,2 S)-diol ( 6) (ring-d 5 and 2,3- 13C 2); in all other metabolites produced, the 2,3- 13C 2 label is washed out. Incubation of the fungus with 4-fluorobenzaldehyde ( 13) produces a pooling of predominantly erythro (1 R,2 S) 1-(4′-fluorophenyl)-1,2-propane diol ( 18 as diacetate) (through the corresponding α-ketols 16, 17). Blocking the para-position with fluorine thus appears to prevent ring oxygenation and also chlorination, forcing the conclusion that para-ring oxygenation precedes meta-chlorination.