Identification of a tomatinase in the tomato-pathogenic actinomycete Clavibacter michiganensis subsp. michiganensis NCPPB382.
The insertion site of a transposon mutant of Clavibacter michiganensis subsp. michiganensis NCPPB382 was cloned and found to be located in the gene tomA encoding a member of the glycosyl hydrolase family 10. The intact gene was obtained from a cosmid library of C. michiganensis subsp. michiganensis. The deduced protein TomA (543 amino acids, 58 kDa) contains a predicted signal peptide and two domains, the N-terminal catalytic domain and a C-terminal fibronectin III-like domain. The closest well-characterized relatives of TomA were tomatinases from fungi involved in the detoxification of the tomato saponin alpha-tomatine which acts as a growth inhibitor. Growth inhibition of C. michiganensis subsp. michiganensis by alpha-tomatine was stronger in the tomA mutants than in the wild type. Tomatinase activity assayed by deglycosylation of alpha-tomatine to tomatidine was demonstrated in concentrated culture supernatants of C. michiganensis subsp. michiganensis. No activity was found with the tomA mutants. However, neither the transposon mutant nor a second mutant constructed by gene disruption was affected in virulence on the tomato cv. Moneymaker.
Kaup
Olaf
O
Department of Genetechnology/Microbiology, University of Bielefeld, Universitaetsstr, Bielefeld, Germany.
Gräfen
Ines
I
Zellermann
Eva-Maria
EM
Eichenlaub
Rudolf
R
Gartemann
Karl-Heinz
KH
D039903:Actinobacteria / Q000201:enzymology* / Q000254:growth & development / Q000472:pathogenicity; D000595:Amino Acid Sequence; D015139:Blotting, Southern; D002240:Carbohydrate Sequence; D002855:Chromatography, Thin Layer; D004251:DNA Transposable Elements; D006026:Glycoside Hydrolases / Q000737:chemistry / Q000378:metabolism*; D018551:Solanum lycopersicum / Q000382:microbiology*; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid; D014053:Tomatine / Q000031:analogs & derivatives / Q000737:chemistry
D004251:DNA Transposable Elements; C484751:alpha-tomatine; D014053:Tomatine; D006026:Glycoside Hydrolases; C099475:tomatinase