Expression, purification and use of the soluble domain of Lactobacillus paracasei beta-fructosidase to optimise production of bioethanol from grass fructans.
Microbial inulinases find application in food, pharmaceutical and biofuel industries. Here, a novel Lactobacillus paracasei beta-fructosidase was overexpressed as truncated cytosolic protein ((t)fosEp) in Escherichia coli. Purified (t)fosEp was thermostable (10-50 degrees C) with a pH optimum of 5; it showed highest affinity for bacterial levan (beta[2-6] linked fructose) followed by nystose, chicory inulin, 1-kestose (beta[2-1] linkages) and sucrose (K(m) values of 0.5, 15, 15.6, 49 and 398 mM, respectively). Hydrolysis of polyfructose moieties in agriculturally-sourced grass juice (GJ) with (t)fosEp resulted in the release of >13 mg/ml more bioavailable fructose than was measured in untreated GJ. Bioethanol yields from fermentation experiments with Brewer's yeast and GJ+(t)fosEp were >25% higher than those achieved using untreated GJ feedstock (36.5[+/-4.3] and 28.2[+/-2.7]mg ethanol/ml, respectively). This constitutes the first specific study of the potential to ferment ethanol from grass juice and the utility of a novel core domain of beta-fructosidase from L. paracasei.
Martel
C M
CM
Institute of Life Science and School of Medicine, Swansea University, Swansea SA2 8PP, Wales, UK.
Warrilow
A G S
AG
Jackson
C J
CJ
Mullins
J G L
JG
Togawa
R C
RC
Parker
J E
JE
Morris
M S
MS
Donnison
I S
IS
Kelly
D E
DE
Kelly
S L
SL
D000595:Amino Acid Sequence; D056804:Biofuels; D004591:Electrophoresis, Polyacrylamide Gel; D000431:Ethanol / Q000378:metabolism*; D005285:Fermentation; D005630:Fructans / Q000378:metabolism*; D006868:Hydrolysis; D007778:Lactobacillus / Q000201:enzymology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D006109:Poaceae / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D012995:Solubility; D013379:Substrate Specificity; D015003:Yeasts / Q000254:growth & development; D043324:beta-Fructofuranosidase / Q000737:chemistry*
D056804:Biofuels; D005630:Fructans; D011994:Recombinant Proteins; D000431:Ethanol; D043324:beta-Fructofuranosidase