A novel purification procedure for chalcone flavanone isomerase from Petunia hybrida and the use of its antibodies to characterize the Po mutation.

We have purified chalcone flavanone isomerase (CHI) from flowerbuds of Petunia hybrida to high purity. We made use of an affinity matrix consisting of Sepharose-bound Dextran Blue that is known to bind proteins containing the dinucleotide fold [S. T. Thompson, K. H. Cass, and E. Stellwagen (1975) Proc. Natl. Acad. Sci. USA 72, 669-672]. The final step, consisting of preparative elution from a denaturing acrylamide gel, yielded an approximately 2000-fold purified CHI protein. The enzyme is a single polypeptide with Mr = 29,000, and highly specific antiserum was raised against it. Using this antiserum it was shown that corolla and anther tissues express different forms of the enzyme as judged by pI. Furthermore, the absence of immunoreactive CHI was demonstrated in a mutant of P. hybrida (genotype popo) which accumulates 2',4,4',6'-tetrahydroxy-chalcone in anthers as a consequence of lack of enzyme activity.