US2021079438A1PendingUtilityA1
Carbohydrate esters as inducers for gene expression
Assignee: QINGDAO VLAND BIOTECH GROUP CO LTDPriority: Jul 20, 2012Filed: Nov 20, 2020Published: Mar 18, 2021
Est. expiryJul 20, 2032(~6 yrs left)· nominal 20-yr term from priority
Inventors:Tom Tao Huang
C07H 13/04C12P 19/12C12P 21/02C12N 9/2411C12N 9/2437C12P 19/44C07H 13/06C12N 9/2408C07H 1/00C07K 14/475C12N 1/14C12N 15/80C12P 21/005C12N 9/2434C12N 1/38C07K 16/00C07H 3/04C07K 14/575
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Claims
Abstract
The invention provides novel carbohydrate esters, in particular disaccharide esters, and the methods of their preparation. These compounds find use as microbial media components for the induction of gene expression in microbial fermentation processes.
Claims
exact text as granted — not AI-modifiedWhat is claimed is:
1 . A method for producing a protein of interest comprising:
a) providing a fermentation host; and b) culturing the fermentation host with a carbon source and one or more compounds selected from the group consisting of:
where R a is H or C(O)R;
R b is H or C(O)R;
R c is H or C(O)R;
R d is H or C(O)CH 3 ;
R e is H or C(O)CH 3 ;
R f is H or C(O)CH 3 ;
R g is H or C(O)CH 3 ; and
R is an aliphatic moiety, with the proviso that in formula (II), R c is H when R a is H or C(O)CH 3 and R b is H or C(O)CH 3 ,
wherein the fermentation host is cultured under conditions sufficient to produce a protein of interest.
2 . The method according to claim 1 , wherein an optional conventional inducer is added to the culture.
3 . The method according to claim 2 , wherein the optional conventional inducer comprises lactose, cellobiose, sophorose, cellulose, cellulose hydrolysate, maltose, isomaltose, maltodextrins, starch, or starch hydrolysate.
4 . The method according to claim 1 , wherein the protein of interest is cellulase or amylase.
5 . The method according to claim 1 , wherein the protein of interest is a homologous or heterologous protein.
6 . The method according to claim 5 , wherein the homologous or heterologous protein is an enzyme, a hormone, a growth factor, a cytokine or an antibody.
7 . The method according to claim 1 , wherein the fermentation host is capable of producing cellulase or amylase.
8 . The method according to claim 1 , wherein the fermentation host is a filamentous fungus or bacteria or other hosts selected from the genus group consisting of Trichoderma, Humicola, Pleurotus, Fusarium, Aspergillus, Streptomyces, Thermomonospora, Bacillus, Cellulomonas, Penicillium, Basidiomycete, Chrysoporium, Pestalotiopsis, Neurospora, Cephalosporium, Achlya, Podospora, Endothia, Mucor, Cochliobolus, Myceliopthora, Talaromyces , and Pyricularia.
9 . The method according to claim 1 , wherein the fermentation host is the filamentous fungus Trichoderma reesei or Aspergillus niger.
10 . The method according to claim 1 , wherein the fermentation host is grown in a liquid culture or on a solid substrate without free-flowing liquid.
11 . The method according to claim 1 , wherein the fermentation host has a promoter operably linked to a gene encoding a protein of interest.
12 . The method according to claim 11 , wherein the promoter is a cellulase gene promoter or an amylase gene promoter.
13 . The method according to claim 11 , wherein the promoter is a cbh1 promoter or a glaA promoter or an amyA promoter.
14 . The method according to claim 1 , wherein the carbon source is biomass.
15 . The method according to claim 14 , wherein the biomass is glucose, sucrose, fructose, glycerol, lactose, cellulose, cellulose hydrolysate, starch, starch hydrolysate, maltose, or maltodextrin.
16 . The method according to claim 1 , wherein the compound of formula (I), (II), (III), (IV) and (V) is isolated from a crude product mixture from either a chemical or enzymatically catalyzed trans-esterification reaction.
17 . The method according to claim 1 , wherein the compounds of formula (VI) are isolated from formic acid treatment process of natural lactonic sophorolipid.
18 . The method according to claim 1 , wherein the compounds of formula (VII) are isolated from the culture of the yeast Candida bombicola.
19 . The method according to claim 1 , wherein R is an aliphatic moiety selected from unsubstituted C 1 -C 24 alkyl, substituted C 1 -C 24 alkyl, unsubstituted C 2 -C 24 alkenyl, and substituted C 2 -C 24 alkenyl.
20 . The method according to claim 1 , wherein R is an aliphatic moiety selected from C 1 -C 24 alkyl substituted with hydroxyl groups and C 2 -C 24 alkenyl substituted with hydroxyl groups.
21 . The method according to claim 1 , wherein R is an aliphatic moiety selected from C 1 -C 24 alkyl substituted with carboxyl groups and C 2 -C 24 alkenyl substituted with carboxyl groups.
22 . The method according to claim 1 , wherein R is an aliphatic moiety selected from C 1 -C 24 alkyl substituted with aromatic groups and C 2 -C 24 alkenyl substituted with aromatic groups.
23 . The method according to claim 1 , wherein the compound of formula (I), (II), (III), (IV), (V), (VI), and (VII) comprises:
where R d is H or C(O)CH 3 ;
R e is H or C(O)CH 3 ;
R f is H or C(O)CH 3 ;
and R g is H or C(O)CH 3 .
24 . A method for producing one or more compounds of formula (I), formula (II), formula (III), formula (IV) or formula (V) comprising:
a) providing a disaccharide; and b) contacting the disaccharide with a vinyl ester in the presence of a lipase enzyme in a solvent system.
25 . The method according to claim 24 , wherein the disaccharide comprises lactose, sophorose, cellobiose, maltose, or isomaltose.
26 . The method according to claim 24 , wherein the lipase enzyme comprises Novozyme 435, Lipozyme TL, Lipozyme RM, or Amano lipase PS.
27 . The method according to claim 24 , wherein the vinyl ester comprises vinyl acetate, vinyl propionate, vinyl butyrate, vinyl cinnamate, vinyl methacrylate, vinyl oleate, vinyl linoleate, vinyl palmitate, or vinyl stearate.
28 . The method according to claim 24 , wherein the solvent system comprises a polar solvent mixed with tert-amyl alcohol, tert-butyl alcohol, tetrahydrofuran, acetone, methanol, or acetonitrile.
29 . The method according to claim 24 , wherein the polar solvent comprises dimethyl sulfoxide (DMSO), pyridine, or dimethylformamide (DMF).
30 . The method according to claim 24 , further comprising heating to a temperature between 40° C. to 100° C.
31 . The method according to claim 24 , further comprising vacuum distillation to remove the solvent system and isolating the product of interest.
32 . A method for producing one or more compounds of formula (I), formula (II), formula (III), formula (IV) or formula (V) by an acid or base catalyzed trans-esterification reaction of an appropriate disaccharide with a suitable vinyl ester in a solvent system.
33 . The method according to claim 32 , wherein the base is potassium carbonate (K 2 CO 3 ), sodium hydroxide (NaOH), or potassium hydroxide (KOH).
34 . The method according to claim 32 , wherein the acid is sulfuric acid (H 2 SO 4 ) or hydrochloric acid (HCl).
35 . The method according to claim 32 , wherein the vinyl ester comprises vinyl acetate, vinyl propionate, vinyl butyrate, vinyl cinnamate, vinyl methacrylate, vinyl oleate, vinyl linoleate, vinyl palmitate, or vinyl stearate.
36 . The method according to claim 32 , wherein the solvent system comprises a polar solvent such as dimethyl sulfoxide (DMSO), pyridine, or dimethylformamide (DMF).
37 . A method for producing one or more compounds of formula (VI) comprising:
a) reacting natural lactonic sophorolipid with formic acid or acetic acid under substantially non-aqueous conditions; and b) partially or completely removing formate or acetate esters formed.
38 . The method according to claim 37 , wherein the reacting natural sophorolipid with formic acid or acetic acid step is catalyzed by a second acid.
39 . The method according to claim 37 , wherein the second acid comprises sulfuric acid (H 2 SO 4 ) or hydrochloric acid (HCl).
40 . The method according to claim 37 , wherein the natural sophorolipid is produced by a fermentation host.
41 . The method according to claim 40 , wherein the fermentation host is Candida bombicola.
42 . The method according to claim 37 , wherein the substantially non-aqueous conditions comprise a reaction mixture having a water content of no more than about 10%.
43 . The method according to claim 37 , further comprising the step of heating at a temperature between 40° C. to 120° C. before the step of partially or completely removing the formate or acetate esters formed.
44 . The method according to claim 37 , wherein removal of the formate esters formed comprises refluxing in acetic acid and methanol.
45 . The method according to claim 37 , wherein removal of the acetate esters formed comprises treating with an acid or a base.
46 . A method for producing one or more compounds of formula (VII) by a fermentation host.
47 . The method according to claim 46 , wherein the fermentation host is Candida bombicola.
48 . The method according to claim 46 , wherein the compounds of formula (VII) and (VIIa) are isolated from a crude mixture from a culture of Candida bombicola.
49 . A method for producing sophorose comprising:
a) reacting natural sophorosides with formic acid under substantially non-aqueous conditions; and b) completely removing formate esters formed;
wherein a purified sophorose is obtained.
50 . The method according to claim 49 , wherein the natural sophorosides are steviosides, sophorolipids, and flavonoid sophorosides.
51 . The method according to claim 49 , wherein the reacting natural sophorosides with formic acid step is catalyzed by a second acid.
52 . The method according to claim 51 , wherein the second acid comprises sulfuric acid (H 2 SO 4 ) or hydrochloric acid (HCl).
53 . The method according to claim 49 , wherein the substantially non-aqueous conditions comprise a reaction mixture having a water content of no more than about 10%.
54 . The method according to claim 49 , further comprising the step of heating at a temperature between 40° C. to 120° C. before the step of completely removing the formate esters.
55 . The method according to claim 49 , wherein completely removing the formate esters comprises treating with a base in a solvent.
56 . The method according to claim 55 , wherein the base is sodium methoxide (CH 3 NaO) or ammonia (NH 3 ) and the solvent is methanol or ethanol.
57 . A compound having a structure of formula (II):
wherein:
R a is H or C(O)R;
R b is H or C(O)R;
R c is H or C(O)R; and
R is an aliphatic moiety, with the proviso that R c is H when R a is H or C(O)CH 3 and R b is H or C(O)CH 3 .
58 . The compound of claim 57 , wherein R is selected from the group consisting of unsubstituted C 1 -C 24 alkyl; C 1 -C 24 alkyl substituted with hydroxyl, carboxyl or aromatic groups;
unsubstituted C 2 -C 24 alkenyl; and C 2 -C 24 alkenyl substituted with hydroxyl, carboxyl or aromatic groups.
59 . The compound of claim 57 , wherein R is an aliphatic moiety selected from unsubstituted C 1 -C 24 alkyl, substituted C 1 -C 24 alkyl, unsubstituted C 2 -C 24 alkenyl, and substituted C 2 -C 24 alkenyl.
60 . The compound of claim 57 , wherein R is an aliphatic moiety selected from C 1 -C 24 alkyl substituted with hydroxyl groups and C 2 -C 24 alkenyl substituted with hydroxyl groups.
61 . The compound of claim 57 , wherein R is an aliphatic moiety selected from C 1 -C 24 alkyl substituted with carboxyl groups and C 2 -C 24 alkenyl substituted with carboxyl groups.
62 . The compound of claim 57 , wherein R is an aliphatic moiety selected from C 1 -C 24 alkyl substituted with aromatic groups and C 2 -C 24 alkenyl substituted with aromatic groups.Cited by (0)
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