Methods and compositions for preparing tagatose from fructose
Inventors
Zanghellini, Alexandre • Roberts, Kyle • COCKREM, Michael Charles Miller • DUNCKLEY, Christopher
Assignees
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Publication Number
US-11060119-B2
Publication Date
2021-07-13
Expiration Date
Abstract
Disclosed herein are compositions and methods for preparing tagatose from fructose, more particularly, compositions comprising thermophilic fructose C4-epimerases derived from thermophilic microorganisms and methods for preparing tagatose from fructose using the compositions.
Core Innovation
The invention relates to a D-fructose C4-epimerase polypeptide (FC4E) and polynucleotides encoding FC4E polypeptides that convert fructose to tagatose through epimerization at the carbon-4 position of fructose. Modified polypeptides are provided that include one or more amino acid substitutions relative to an unmodified polypeptide sequence identified as SEQ ID NO:6, with specified substitution positions and sequence-identity constraints.
The disclosed modified polypeptides are at least 85% identical to SEQ ID NO:6 and include amino acid substitutions at residues corresponding to position H45, M212, or Y340. The modified polypeptides also comprise H at position 85, D at position 121, H at position 247, D at position 250, and G at position 271. In addition to the polypeptide embodiments, the disclosure covers polynucleotides encoding the modified polypeptides and expression vectors/promoters for expressing these polypeptides in microbial or higher cells.
The invention further addresses production and processing of tagatose from fructose using the FC4E polypeptides and/or microorganisms genetically modified to express them. Embodiments include immobilization of the polypeptides or microorganisms on organic or inorganic carriers/supports, isolating tagatose from a feed using chromatographic methods, including simulated moving bed chromatography, and crystallization of the tagatose product.
Claims Coverage
The independent claims cover one modified-polypeptide inventive core centered on SEQ ID NO:6, with specified substitution positions, sequence identity, and residue constraints. Related claim material also addresses tagatose production and processing using the polypeptide or a microorganism expressing it.
Modified polypeptide for fructose-to-tagatose carbon-4 epimerization
A modified polypeptide comprising one or more amino acid substitutions in an unmodified polypeptide, wherein the amino acid sequence of the unmodified polypeptide is SEQ ID NO:6 and wherein at least one substituted amino acid is at the amino acid residues corresponding to position H45, M212, or Y340, wherein the modified polypeptide converts fructose to tagatose through epimerization at the carbon-4 position of fructose.
Sequence identity and specified residue constraints relative to SEQ ID NO:6
The modified polypeptide is at least 85% identical to SEQ ID NO:6, and comprises H at position 85, D at position 121, H at position 247, D at position 250, and G at position 271.
The claim coverage centers on a SEQ ID NO:6-based FC4E modified polypeptide with specified substitution positions and sequence-identity criteria that converts fructose to tagatose via carbon-4 epimerization. Related claim material also includes contacting fructose with the polypeptide or a microorganism expressing it, immobilization, chromatographic isolation, and crystallization embodiments.
Stated Advantages
Not explicitly described in patent.
Documented Applications
Producing a tagatose composition by contacting a fructose-containing starting composition with a polypeptide or a microorganism expressing the polypeptide.
Isolating tagatose from a feed using a chromatographic method.
Crystallizing tagatose to obtain high purity tagatose crystals.
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