Publications

• The Fusarium solani gene encoding kievitone hydratase, a secreted enzyme that catalyzes detoxification of a bean phytoalexin.

  Li D., Chung K.R., Smith D.A., Schardl C.L.

  Among the antimicrobial phytoalexins produced by Phaseolus vulgaris (French bean) is the prenylated isoflavonoid, kievitone. The bean pathogen, Fusarium solani f. sp. phaseoli, secretes a glycoenzyme, kievitone hydratase (EC 4.2.1.95), which catalyzes conversion of kievitone to a less toxic metabo ... >> More

  Among the antimicrobial phytoalexins produced by Phaseolus vulgaris (French bean) is the prenylated isoflavonoid, kievitone. The bean pathogen, Fusarium solani f. sp. phaseoli, secretes a glycoenzyme, kievitone hydratase (EC 4.2.1.95), which catalyzes conversion of kievitone to a less toxic metabolite. Among F. solani strains, those that are highly virulent to P. vulgaris also produce kievitone hydratase constitutively, suggesting that the enzyme is a virulence factor. Based on the N-terminal amino acid sequence of purified enzyme, the kievitone hydratase cDNA and gene (khs) were cloned. The identities of khs and the cDNA were confirmed by their expression in transgenic Neurospora crassa and Emericella nidulans. Based on the gene and cDNA sequences, khs is predicted to encode a preprotein of 350 amino acids, from which a 19 amino acid N-terminal transit peptide is removed during maturation and secretion. The predicted mass of the mature polypeptide, 37 kDa, contrasts with the 47 to 49 kDa size estimated by electrophoresis of purified enzyme, confirming that the enzyme is extensively glycosylated. The inferred polypeptide sequence has seven canonical sites for N-glycosylation. Southern blot-hybridization analysis of F. s. f. sp. phaseoli DNA indicates one khs locus and an additional locus with weak hybridization to the khs probe. Sequences related to khs were also detected in several isolates of F. solani and the related teleomorph, Nectria haematococca. However, strains of F. oxysporum known to exhibit inducible kievitone hydratase activity (but not pathogenic to bean) did not have detectable khs homology. Nevertheless, all isolates known to cause severe disease on bean possessed khs sequence. << Less

  Mol. Plant Microbe Interact. 8:388-397(1995) [PubMed] [EuropePMC]

• Induction and purification of kievitone hydratase from Fusarium solani f. sp. phaseoli.

  Turbek C.S., Li D.X., Choi G.H., Schardl C.L., Smith D.A.

  Fusarium solani f. sp. phaseoli is capable of detoxifying the major isoflavonoid phytoalexins produced by its host plant Phaseolus vulgaris. One of the enzymic activities involved is kievitone hydratase (KHase), a secreted glycoprotein which catalyses the conversion of kievitone to the less fungit ... >> More

  Fusarium solani f. sp. phaseoli is capable of detoxifying the major isoflavonoid phytoalexins produced by its host plant Phaseolus vulgaris. One of the enzymic activities involved is kievitone hydratase (KHase), a secreted glycoprotein which catalyses the conversion of kievitone to the less fungitoxic derivative, kievitone hydrate. Even under conditions of substrate induction, the enzyme is expressed at levels that are too low for satisfactory purification. Therefore, several other isoflavonoids were tested as inducers in culture. Among the phytoalexins produced by the host plant, phaseollinisoflavan was the best inducer, elevating the level of secreted enzyme eight-fold. Treatment with biochanin A, a product of chickpea, resulted in a 16-fold increase of secreted activity. The maximum rate of induction was observed 9-24 hr after addition of biochanin A, during which time several metabolites of the inducer were also present. KHase was purified from filtrates of biochanin A-induced cultures. Denaturing gel electrophoresis indicated that two species of Mr 47,000 and 49,000 copurified with the activity. N-Terminal sequence analysis indicated that the two species possessed related, or identical, polypeptide moieties. Comparison with the size of the non-denatured enzyme, previously determined to be ca 100,000, indicates that its native state is a dimer. << Less

  Phytochemistry 29:2841-2846(1990) [PubMed] [EuropePMC]