Purification and characterization of beta(2)-tomatinase, an enzyme involved in the degradation of alpha-tomatine and isolation of the gene encoding beta(2)-tomatinase from Septoria lycopersici

Lycopersicon species often contain the toxic glycoalkaloid alpha-tomatine, which is proposed to protect these plants from general microbial infection, However, fungal pathogens of tomato often are tolerant to alpha-tomatine and detoxification of alpha-tomatine may be how these pathogens avoid this potential barrier, As an initial step to evaluate this possibility, we have purified to homogeneity a beta-1,2-D glucosidase from the tomato pathogen Septoria lycopersici that hydrolyzes the beta-1,2-D glucosyl bond on the tetrasaccharide moiety of alpha-tomatine to produce beta(2)-tomatine. The enzyme is a 110-kDa protein with a pI of 4.5 and a K-m for alpha-tomatine of 62 mu M, Little or no activity was detected on a variety of other glycosides. The gene encoding this protein was isolated and contains an open reading frame of 803 amino acids that shares sequence homology with several other beta-D-glucosidases. When S. lycopersici was incubated with alpha-tomatine, beta(2)-tomatinase mRNA accumulated, suggesting that the enzyme is substrate inducible, Aspergillus nidulans expressed ''beta(2)-tomatinase'' activity when transformed with this gene but transformants were still sensitive to alpha-tomatine.