Identification and characterization of a glucan-producing enzyme from Lactobacillus hilgardii TMW 1.828 involved in granule formation of water kefir.
Water kefir is a home made fermented beverage based on a sucrose solution with fruit extracts. The inoculum of such fermentations consists of macroscopic granula containing lactic and acetic acid bacteria, and yeasts, which are embedded in an exopolysaccharide (EPS) matrix. In this work, a strain of Lactobacillus hilgardii producing large amounts of the granule-forming dextran could be isolated. The glycosyltransferans (Gtf) commonly called glucansucrase responsible for the production of this dextran was purified from L. hilgardii. Characteristic enzyme kinetic data were obtained. Optimum activity was observed between pH 4.3 and 4.6 and temperatures between 40 degrees C and 45 degrees C. A Michaelis-Menten kinetic could be fit to the experimental data and a K(M) of 0.0385 M was calculated. The corresponding gtf gene was identified and characterized. It encodes a 1448 amino acid protein with higher homologies to Gtfs of Lactobacillus parabuchneri, Lactobacillus sakei and Lactobacillus fermentum followed by lower homologies to Lactobacillus reuteri Gtfs. By knockout experiments the role of this gene in granule dextran production was demonstrated.
Waldherr
Florian W
FW
Technische Universität München, Technische Mikrobiologie, Weihenstephaner Steig 16, D-85350 Freising, Germany.
Doll
Viktoria M
VM
Meissner
Daniel
D
Vogel
Rudi F
RF
D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D043302:Cultured Milk Products / Q000382:microbiology*; D004795:Enzyme Stability; D005936:Glucans / Q000378:metabolism*; D016695:Glycosyltransferases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D007778:Lactobacillus / Q000145:classification / Q000201:enzymology* / Q000235:genetics / Q000302:isolation & purification*; D010802:Phylogeny
D001426:Bacterial Proteins; D005936:Glucans; D016695:Glycosyltransferases; C033837:alternansucrase