The biosynthesis of cyanogenic glucosides in higher plants. Identification of three hydroxylation steps in the biosynthesis of dhurrin in Sorghum bicolor (L.) Moench and the involvement of 1-ACI-nitro-2-(p-hydroxyphenyl)ethane as an intermediate.
N-Hydroxytyrosine, (E)- and (Z)-p-hydroxyphenyl-acetaldehyde oxime, p-hydroxyphenylacetonitrile, and p-hydroxymandelonitrile are established intermediates in the biosynthesis of the tyrosine-derived cyanogenic glucoside dhurrin (Halkier, B. A., Olsen, C. E., and Møller, B. L. (1989) J. Biol. Chem. 264, 19487-19494. Simultaneous measurements of oxygen consumption and biosynthetic activity using a microsomal enzyme system isolated from etiolated sorghum seedlings demonstrate a requirement for three oxygen molecules in the conversion of tyrosine to p-hydroxymandelonitrile. Two oxygen molecules are consumed in the conversion of tyrosine to (E)-p-hydroxyphenylacetaldehyde oxime, indicating the existence of a previously undetected hydroxylation step in addition to that resulting in the formation of N-hydroxytyrosine. Radioactively labeled 1-nitro-2-(p-hydroxyphenyl)ethane was chemically synthesized and tested as a possible intermediate. Biosynthetic experiments demonstrate that the microsomal enzyme system metabolizes the nitro compound to the subsequent intermediates in dhurrin synthesis (Km = 0.05 mM; Vmax = 14 nmol/mg of protein/h). Low amounts of 1-nitro-2-(p-hydroxyphenyl)ethane are produced in the microsomal reaction mixtures when tyrosine is used as substrate. These data support the involvement of 1-nitro-2-(p-hydroxyphenyl)ethane or more likely its aci-nitro tautomer as an intermediate between N-hydroxytyrosine and p-hydroxyphenylacetaldehyde oxime. The conversion of (E)-p-hydroxyphenylacetaldehydeoxime to p-hydroxymandelonitrile requires a single oxygen molecule. The oxygen molecule is utilized for hydroxylation of p-hydroxyphenylacetonitrile into p-hydroxymandelonitrile. This indicates that the conversion of p-hydroxyphenylacetaldehyde oxime into p-hydroxyphenylacetonitrile proceeds by a simple dehydration reaction.
Halkier
B A
BA
Department of Plant Biology, Royal Veterinary and Agricultural University, Copenhagen, Denmark.
Møller
B L
BL
D002250:Carbon Radioisotopes; D005960:Glucosides / Q000096:biosynthesis*; D006863:Hydrogen-Ion Concentration; D006900:Hydroxylation; D007700:Kinetics; D008861:Microsomes / Q000378:metabolism; D009570:Nitriles / Q000378:metabolism*; D009595:Nitroparaffins / Q000378:metabolism*; D010101:Oxygen Consumption; D010944:Plants / Q000378:metabolism*; D014316:Tritium; D014443:Tyrosine / Q000378:metabolism
D002250:Carbon Radioisotopes; D005960:Glucosides; D009570:Nitriles; D009595:Nitroparaffins; D014316:Tritium; C066540:1-nitro-2-(4-hydroxyphenyl)ethane; D014443:Tyrosine; C011220:dhurrin