Dx. Li et al., THE FUSARIUM-SOLANI GENE ENCODING KIEVITONE HYDRATASE, A SECRETED ENZYME THAT CATALYZES DETOXIFICATION OF A BEAN PHYTOALEXIN, Molecular plant-microbe interactions, 8(3), 1995, pp. 388-397
Among the antimicrobial phytoalexins produced by Phaseolus vulgaris (F
rench bean) is the prenylated isoflavonoid, kievitone. The bean pathog
en, 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 constitutive
ly, suggesting that the enzyme is a virulence factor. Based on the N-t
erminal amino acid sequence of purified enzyme, the kievitone hydratas
e 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 pre
dicted to encode a preprotein of 350 amino acids, from which a 19 amin
o acid N-terminal transit peptide is removed during maturation and sec
retion. The predicted mass of the mature polypeptide, 37 kDa, contrast
s with the 47 to 49 kDa size estimated by electrophoresis of purified
enzyme, confirming that the enzyme is extensively glycosylated. The in
ferred polypeptide sequence has seven canonical sites for N-glycosylat
ion. Southern blot-hybridization analysis of F. s. f. sp. phaseoli DNA
indicates one khs locus and an additonal locus with weak hybridizatio
n to the khs probe. Sequences related to khs were also detected in sev
eral isolates of F. solani and the related teleomorph, Nectria haemato
cocca. However, strains of F. oxysporum known to exhibit inducible kie
vitone hydratase activity (but not pathogenic to bean) did not have de
tectable khs homology. Nevertheless, all isolates known to cause sever
e disease on bean possessed khs sequence.