Medicarpin, the major phytoalexin in alfalfa, is synthesized by way of the 
isoflavonoid branch of phenylpropanoid metabolism. One of the final steps of 
medicarpin biosynthesis, from vestitone to 7,2'-dihydroxy-4'-methoxyisoflavanol, 
is catalyzed by vestitone reductase. A 1245-bp cDNA clone which encodes 
vestitone reductase was identified utilizing internal amino acid sequence of 
purified vestitone reductase. When expressed in Escherichia coli, the cloned 
enzyme exhibits strict substrate stereospecificity for (3R)-vestitone, as was 
observed for vestitone reductase purified from alfalfa. The calculated molecular 
weight of the protein (35,918) is similar to that of purified vestitone 
reductase from alfalfa (38 kDa by SDS-PAGE). The levels of vestitone reductase 
transcript (1.35 kb) greatly increase within 2 h of elicitor addition to alfalfa 
cell suspension cultures, preceding the rapid increases in vestitione reductase 
enzyme activity and medicarpin biosynthesis. In healthy alfalfa plants, the 
highest levels of transcripts were detected in roots and root nodules, 
consistent with the synthesis of medicarpin and its conjugate in these tissues. 
The cloning of the vestitone reductase gene provides a specific tool for the 
study and manipulation of pterocarpan biosynthesis in legumes.