KR20190120779A - Polyamine high yeast and food and drink composition comprising the same - Google Patents

Abstract

The present invention, in comparison with the association No. 7 yeast, amino acid sequence of any one or more of leucine synthase, aconitate synthase, S-adenosyl-L-methionine synthase, aspartic acid / glutamic acid transport protein, The LEU1 gene encoded by the one or several amino acids substituted, deleted, inserted or added, or a combination of two or more of these, or the third chromosome, the ACO2 gene encoded by the tenth chromosome, twelfth In one or more base acid sequences of the SAM1 gene encoded on the chromosome or the AGC1 gene encoded on the 16th chromosome, one or several bases are substituted, deleted, inserted or added, or two of them The yeast variant which is mutated by the above combination and has high polyamine is provided.

Description

Polyamine high yeast and food and drink composition comprising the same

The present invention relates to high polyamine-containing yeast, especially high spermidine-containing yeast, and a food and drink composition comprising the same.

In the field of functional food, it is required to be able to activate cells. Among them, a substance having a high effect of a linear in vivo aliphatic hydrocarbon having two or more primary amino groups called polyamines is provided as food. Representative polyamines include putrescine, spermidine and spermine. Physiological actions of polyamines include (1) cell proliferation, (2) cell differentiation promotion, (3) immune essential factors, (4) antiallergic action, (5) protein synthesis promotion, and (6) nucleic acid Stabilization of the structure by interaction, (7) enzyme activity control action and the like are known. Recently, it has been reported that lifespan is extended by orally ingested polyamines.

As a manufacturing method of the polyamine or polyamine composition which can be used industrially, the method of processing and preparing a yeast cell or yeast culture liquid under acidic conditions (patent document 1) is disclosed. In addition, as a method of preparing a polyamine composition from a plant material, a method of extracting a plant material by treating it under acidic conditions (Patent Document 2), and adding a salt solution such as salt, magnesium chloride, and calcium chloride to the plant material to obtain a polyamine extract. The manufacturing method (patent document 3) has been examined. In fact, as a food containing high polyamine, there are rice extract, soybean extract and yeast extract. Although these polyamine-rich foods are known to activate cells, foods that activate cells are required. In addition, a method of obtaining a high polyamine-containing yeast by mixing a polyamine as a reagent in a culture medium of a yeast is also known (Patent Document 4). However, the production of a polyamine is not produced by the yeast itself, and a pure product of a polyamine which is not recognized as a food is externally obtained. Since the method of introduction is taken, there is a problem that providing such a yeast as a food is not recognized. In addition, it is possible to increase the concentration of polyamines contained in the cells by applying stress (penetration pressure due to ethanol addition, salt addition, etc.) when culturing yeast (Non-Patent Document 1), but growth of yeast is inhibited by stress. Therefore, there is a problem that productivity is extremely low.

Among the polyamines, spermidine and spermine have high body utilization and are excellent in terms of functionality. In particular, spermidine has a higher allowable daily intake than spermine and is preferable to be consumed in a larger amount (Non-Patent Document 2).

Japanese Patent Application Laid-Open No. H10-52291 Japanese Patent Application Laid-Open No. H10-101624 Japanese Patent Publication No. 2010-263816 Japanese Patent Application Laid-Open No. 2016-187336

Japan Agricultural Chemistry Society 1995 Conference Abstracts Collection p.129 Food Chem Toxcol, 35 (3) 337-348 (1997)

An object of the present invention is to provide a polyamine high yeast, particularly spermidine high yeast, which activates cells more, and a food and drink composition comprising the same.

Conventionally, the method of obtaining a polyamine high food content as an extract has been used by extracting a polyamine with an acid etc. from the plant material high polyamine content, and concentrating. As a result of studying the problem-solving eagerly, the present inventor has unexpectedly succeeded in producing a mutant yeast containing high polyamine without introducing a polyamine which is a reagent from the outside into the yeast and stressing a high concentration salt, etc. The present invention has been completed. Since the yeast according to the present invention contains a high amount of polyamine in the cells, the polyamine accumulates at high concentration in the yeast cells without going through the extraction and concentration of the polyamine.

That is, this invention includes the following inventions.

[One]

In comparison with Association No. 7 yeast,

In any one or more amino acid sequence of leucine synthase, aconitate synthase, S-adenosyl-L-methionine synthase, or aspartic acid / glutamic acid transport protein, one or several amino acids are substituted, deleted, inserted Or in addition, or in combination of two or more of these, or

One or more nucleotide sequences of the LEU1 gene encoded on the third chromosome, the ACO2 gene encoded on the chromosome 10, the SAM1 gene encoded on the chromosome 12, and the AGC1 gene encoded on the chromosome 16 Wherein one or several bases are substituted, deleted, inserted or added, or mutated to two or more of these,

Furthermore, yeast variant which has high polyamine.

[2]

Any one of the bases at the positions corresponding to the 266th C of the LEU1 gene, the 1964th T of the ACO2 gene, the 1130th C of the SAM1 gene, and the 744th A of the AGC1 gene in association 7 yeast Or the yeast variant of [1] in which more than one base is substituted.

[3]

The base corresponding to the 266th C of the LEU1 gene is A, and the base corresponding to the 1964th T of the ACO2 gene, which is encoded on the 10th chromosome, is C, and the 1130th C of the SAM1 gene, which is encoded on the 12th chromosome The yeast variant of [2] whose corresponding base is G and the base corresponding to 744th A of AGC1 gene encoded by the 16th chromosome is T.

[4]

The yeast variant in any one of [1]-[3] which is a yeast which belongs to Saccharomyces.

[5]

The yeast variant in any one of [1]-[4] whose content of polyamine is a total amount of putrescine, spermidine, and spermine.

[6]

The food-drinks composition containing the yeast variant and crystalline polysaccharide in any one of [1]-[5].

[7]

The composition as described in [6] whose weight ratio of a yeast variant and a crystalline polysaccharide is 1: 0.1-1.

[8]

The composition as described in [6] or [7] whose crystalline polysaccharide is cyclodextrin and / or indigestible dextrin.

[9]

In comparison with Association No. 7 yeast,

In any one or more amino acid sequence of leucine synthase, aconitate synthase, S-adenosyl-L-methionine synthase, or aspartic acid / glutamic acid transport protein, one or several amino acids are substituted, deleted, inserted Or in addition, or in combination with two or more of these, or

One or more nucleotide sequences of the LEU1 gene encoded on the third chromosome, the ACO2 gene encoded on the chromosome 10, the SAM1 gene encoded on the chromosome 12, and the AGC1 gene encoded on the chromosome 16 Wherein one or several bases are substituted, deleted, inserted or added, or mutated to a combination of two or more thereof,

A method for producing a yeast variant having high polyamine.

The genome of the resulting mutant strain was analyzed using Illumina HiSeq, and compared with the arrangement of wild strains. As a result, homogeneity of four genes related to the synthesis of polyamines was confirmed. The 266th C of the leucine synthetase LEU1 encoded on the third chromosome is A, the 1964th T of the aconitic acid synthetic gene ACO2 encoded on the 10th chromosome is C, and the S is encoded on the 12th chromosome. The 1130th C of the adenosyl-L-methionine synthetic gene SAM1 was changed to G, and the 744th A of the aspartic acid / glutamic acid transport gene AGC1 encoded on the 16th chromosome was changed to T. Arginase gene CAR1 coded for chromosome 16, ornithintal carbonate gene SPE1 coded for chromosome 11, spermidine synthetic gene SPE3 coded for chromosome 16, as a polyamine synthetic gene Although the spermine synthetic gene SPE4 encoded by the chromosome is known, it has been found that a high polyamine production line is obtained by mutation of genes other than these. In addition, since the polyamine high producer strain according to the present invention was not produced by genetic modification, it is considered to be suitable as a food.

Therefore, according to the present invention, it is possible to provide a polyamine high yeast, particularly spermidine high yeast, which is effective for cell activation, and food and drink containing the same. Yeast according to the present invention can exert the effect of cell activation from conventional rice extract, soybean extract and yeast extract of the same polyamine content by containing polyamine in yeast. As a secondary effect, it has been clarified that the polyamine high yeast obtained in the present invention has excellent thermal stability of polyamines including spermidine when combined with crystalline polysaccharides. For this reason, by combining the yeast of this invention with a crystalline polysaccharide, it becomes possible to provide the polyamine high food-drinks which are easy to heat-cook.

1 shows the nucleotide sequence of the LEU1 gene of yeast according to the present invention compared to wild type (association No. 7 yeast).
Figure 2a shows the base sequence of the ACO2 gene of yeast according to the present invention compared to wild type (Association No. 7 yeast).
Figure 2b shows the nucleotide sequence of the ACO2 gene of yeast according to the present invention compared to wild type (association No. 7 yeast).
Figure 3 shows the base sequence of the SAM1 gene of yeast according to the present invention compared to wild type (association No. 7 yeast).
Figure 4a shows the base sequence of the AGC1 gene of yeast according to the present invention compared to wild type (association No. 7 yeast).
4b shows the nucleotide sequence of the AGC1 gene of yeast according to the present invention compared to wild type (Association No. 7 yeast).

(High polyamine yeast)

In one embodiment, the present invention provides one or more amino acids in leucine synthetase, aconitic acid synthase, S-adenosyl-L-methionine synthase, aspartic acid / glutamic acid transport protein as compared to Association No. 7 yeast. In an arrangement, up to 100, preferably up to 20, more preferably one or several, for example two, three, four, five, six, seven, eight, nine Or 10 amino acids are mutated, substituted, deleted, inserted or added, or a combination of two or more thereof, or

One or more nucleotide sequences of the LEU1 gene encoded on the third chromosome, the ACO2 gene encoded on the chromosome 10, the SAM1 gene encoded on the chromosome 12, and the AGC1 gene encoded on the chromosome 16 Wherein one or several, for example two, three, four, five, six, seven, eight, nine or ten bases are substituted, deleted, inserted or added, or two of these Mutations in more than one combination,

Moreover, the yeast variant (henceforth "variant yeast", a "variant", and "polyamine high content yeast") which has high polyamine is provided.

More specifically, the yeast variant according to the present invention may contain at least 1.5 mg, preferably at least 2.0 mg, more preferably at least 3.0 mg, even more preferably at least 10 mg of polyamine per 1 g (dry weight) of yeast. .

Mutant yeast can be obtained by mutating yeast belonging to the genus Saccharomyces, preferably Saccharomyces cerevisiae by methods known to those skilled in the art. More specifically, the mutant of the present invention is obtained by irradiating ultraviolet light to the Australian Brewing Association 7 (hereinafter also referred to simply as "Association 7"). The variant of the present invention can be obtained by irradiating ultraviolet rays to Saccharomyces cerevisiae IFO2346 described in Japanese Patent No. 5974891.

Or in order to produce the yeast containing a high polyamine, you may start from another state with high ability to synthesize | combine a polyamine. Although it is possible to cultivate yeast in a medium containing a large amount of polyamine or to apply high stress to the polyamine contained in the cells by applying stress (penetration pressure due to ethanol addition, salt addition, etc.), such a variant may perform such manipulation. It differs from the conventional yeast in that it has a function of producing a large amount of polyamine without accumulating and accumulating in cells. In the case of culturing while imparting infiltration pressure stress due to ethanol, salt, or the like, since the growth of yeast is inhibited by the influence of stress, productivity is extremely low. However, in the case of such a variant, high polyamine-containing yeast such as spermidine-containing yeast is obtained even in culture under a non-stress, and thus the productivity is excellent.

Association 7 used in the Example to obtain a high polyamine-containing yeast is one type of Cheongju yeast (Saccharomyces cerevisiae), and is registered under 101557 in the NBRC (NITE Biological Resource Center).

The high polyamine yeast of the present invention is obtained by culturing yeast as a starting material, such as Association 7, in a medium containing yeast extract, collecting after culturing, heat sterilization, and then spray drying. Alternatively, the polyamine-containing yeast of the present invention is cultured in a medium containing a yeast extract in a yeast extract mutated by a method known to those skilled in the art, such as ultraviolet irradiation, collected after culturing, heat sterilization, and then spray drying It is obtained by adding. The concentration of yeast extract in the medium is preferably 0.1% or more and 10% or less.

The mutation treatment method is not particularly limited, and a known method can be used. For example, chemical treatment in ethyl methanesulfonic acid, N-methyl-N-nitro-N-nitrosoguanidine, nitrous acid, acridine-based dyes and the like; Ultraviolet irradiation; Methods such as irradiation.

Mutation sites may be mutated so that at least one amino acid sequence or base sequence of leucine synthetase, aconitic acid synthase, S-adenosyl-L-methionine synthetase, or aspartic acid / glutamic acid transport protein is high in polyamine. It is not specifically limited. More specifically, 266th C of the LEU1 gene encoded on the 3rd chromosome is A, and 1964th T of the ACO2 gene encoded on the 10th chromosome is C, as compared with the gene sequence of association 7 yeast. It is preferable that the 1130th C of the SAM1 gene encoded on the twelfth chromosome is substituted with G, and the 744th A of the AGC1 gene encoded on the 16th chromosome is substituted with T. Introduction of such a point mutation can also be performed by a well-known genetic engineering technique.

In the present specification, "point variation" refers to monobasic substitution shown on a corresponding base sequence in comparison with the base sequence of Association 7. The point mutation usually includes a transition mutation that is a substitution between purine or pyrimidine, or a transversion variation that is a substitution between purine and pyrimidine. However, in the present invention, any variation may be employed as long as the poly yeast contains a high polyamine. Point mutations are known biological techniques, such as PCR using primers designed to introduce desired mutations, or donors (eg, single-chain oligonucleotides or plasmids) that are homologous to the locus on either side to introduce mutations. Can be introduced according to the genome editing technology (CRISPR / Cas9).

Polyamine is a generic term for linear aliphatic hydrocarbons having a plurality of amino groups bonded thereto. Examples thereof include putrescine, spermidine, spermine, 1,3-diaminopropane, cadaverine, caldine, homospermidine, and aminopropylcarda. Berin, teremin, termospermine, cannabalmin, aminopentylnorspermidine, N, N-bis (aminopropyl) cardaberine, homospermine, caldopentamine, homocaldopentamine, caldohexamine, Homo caldo hexamine spermidine etc. are mentioned. Unless otherwise specified, the term "polyamine" as used herein means one or a plurality of compounds selected from the group consisting of putrescine, spermidine and spermine having high cell activation ability.

The polyamine amount can be measured by summing the amounts of putrescine, spermidine and spermine extracted from the spray-dried yeast. Dry yeast is obtained by taking out a cell from a medium containing yeast by filtration or a centrifuge and drying it. In order to remove a media component, you may wash | clean. Meanwhile, spray drying, reduced pressure drying, lyophilization, or the like may be used for drying the yeast, and those skilled in the art may appropriately select a drying method according to the purpose.

(Composition comprising high polyamine yeast)

In another aspect, the present invention provides a composition comprising a polyamine high yeast, in particular a spermidine high yeast, in particular a composition that activates cells. From the viewpoint of imparting functions such as thermal stability to the polyamine, the composition may contain crystalline polysaccharides in addition to yeast. Crystalline polysaccharide means a polysaccharide that exists naturally as a crystalline solid, and examples thereof include starch, dextrin, glycogen and cellulose. When ordinary saccharides other than crystalline polysaccharides are used in combination with the composition, the saccharides change into a candy form and become hard, resulting in difficulty in handling the composition.

Cyclodextrin which is dextrin which has a cyclic structure can be used suitably as a crystalline polysaccharide. Cyclodextrins have alpha, beta, and gamma forms, and any can be used as the crystalline polysaccharide. Among these, alphacyclodextrin and gammacyclodextrin are preferable. Other dextrins, such as indigestible dextrins, can also be used in the compositions of the present invention.

It is preferable that the weight ratio of the high polyamine dry yeast and crystalline polysaccharide is 1: 0.1-3. More preferably, they are the weight ratio 1: 0.1-1 of polyamine high content yeast and crystalline polysaccharide. Less crystalline polysaccharides lower the stability of the yeast. Moreover, when it is higher than this range, content of polyamine, especially spermidine per composition becomes low too much, and in order to exhibit desired function, such as spermidine in a human body, it is necessary to ingest a composition in large quantities.

By combining polyamine high yeast and crystalline polysaccharide, yeast can be stabilized and handled easily. Mixing of yeast and crystalline polysaccharides may take either dry or wet. When wet, the sugars can be sterilized, mixed with yeast, and dried. Moreover, it is possible to make a composition by mixing, after mixing as undried yeast, without mixing as dry yeast.

Although not intended to be limiting, the composition containing the crystalline polysaccharide is, for example, cultured and concentrated by centrifugation after culturing the yeast for 4 days, and then added γ-cyclodextrin to it at 60 ° C. It is obtained by heating for a time and spray drying this.

It is assumed that the composition of the present invention is used for various purposes. For example, the composition of the present invention, as well as a variety of general foods, food and beverages, such as specific foods that are functionally marked, such that the individual in need of nutritional enhancement, in particular those requiring the activity of cells can be consumed Health foods, nutritional foods, functional skin foods, health foods, special purpose foods, nutritional foods, health foods, supplements, beauty foods, other health foods, or medicines , Quasi-drugs, and cosmetics.

The modified yeast as an active ingredient may be contained in the composition in a state as it is, and may be added to the composition by being crushed and powdered. Additives, such as another bioactive component and excipient, can also be added to a composition.

The form of the composition is also not particularly limited, and may be powder, granule, tablet, capsule, jelly phase, or the like. You may process further about a specific form, for example, may coat the surface of the tablet-like composition obtained by compression tableting. In addition, the granulated powder may be granulated, or the powder or granulated granules may be encapsulated.

The composition of the present invention can be used as food in tablet or powder form. It can also be added to nutritional supplements such as food and beverages or supplements such as juices and dairy products for the purpose of nutritional enhancement. The form may be a commercial form such as a liquid, a tablet, a powder, or a capsule. As another aspect, the composition of the present invention may be used as a seasoning added to food or drink, even if the food or beverage is not itself. It is preferable that 50 to 100% of yeast are contained in a composition.

(Manufacturing method of yeast containing high polyamine)

In another aspect, the present invention provides a method for producing a polyamine high yeast. The manufacturing method of the polyamine high yeast is compared with the association No. 7 yeast,

In any one or more amino acid sequence of leucine synthase, aconitate synthase, S-adenosyl-L-methionine synthase, or aspartic acid / glutamic acid transport protein, one or several amino acids are substituted, deleted, inserted Or in addition, or in combination with two or more of these, or

One or more nucleotide sequences of the LEU1 gene encoded on the third chromosome, the ACO2 gene encoded on the chromosome 10, the SAM1 gene encoded on the chromosome 12, and the AGC1 gene encoded on the chromosome 16 Wherein one or several bases are substituted, deleted, inserted or added, or mutated to a combination of two or more thereof. The method may further include any step of having a polyamine high in the yeast.

Hereinafter, the present invention will be described in more detail with reference to Examples, but the present invention is not limited to these Examples.

Example 1

How to find the polyamine content:

To 10-100 mg of the sample, 4 ml of an aqueous solution of 5% perchloric acid and 2 ppm of 1,7-diaminoheptane was added and suspended, and the polyamine in the powder was extracted by maintaining the mixture at 50 ° C for 10 minutes to 1 hour. Thereafter, the extract was centrifuged, and 0.4 ml of an aqueous solution of 2% perchloric acid and 2 ppm 1,7-diaminoheptane was added to 0.1 ml of the obtained supernatant, followed by mixing in 0.25 ml of 18% aqueous sodium carbonate solution. The mixture was further mixed with 0.5 ml of 1% monoethyl chloride and kept at 45 ° C. for 1 hour. 0.25 ml of 10% aqueous proline solution was added thereto, and the mixture was kept at 45 ° C for 10 minutes. Thereafter, 2 ml of toluene was added to the mixture, and vigorously mixed, 1 ml of the toluene layer was obtained. The toluene layer was further dried by blowing with nitrogen, and redissolved in 0.5 ml of acetonitrile. The result was analyzed by liquid chromatography. The calibration curve sets the concentration of 1,7-diaminoheptane to 2 ppm, and then prepares a dilution series of 0.125 to 8 ppm for each polyamine of spermidine, spermine, and putrescine, and analyzes the liquid chromatograph. It was created from the (peak area value). Unless otherwise specified, the above reagent was used as Wako Pure Chemicals.

Liquid chromatography conditions:

Equipment: Shimadzu Corporation, High Performance Liquid Chromatography, LC-20A

Column: YMC-Pack ODS-TMS (5 μm), 150 × 4.6 mm I.D.

Measurement temperature: 40 ℃

Detection: 510 nm fluorescence in fluorescence detector excitation 365 nm

Eluent: acetonitrile: water = 72: 28

Elution rate: 1.5mL / min

(Acquisition of variation)

As a parent strain before mutation, yeast used Saccharomyces (Association 7). The parent strain was shaken at 30 ° C. for 16 hours in a liquid medium containing 1% Bacto yeast extract, 2% Bacto peptone, and 2% glucose, and the culture medium was irradiated with 45 W of ultraviolet light. Thereafter, the culture solution was diluted 1-100,000 times with 50 mM potassium phosphate buffer (pH7) and applied to an agar medium containing 1% Bacto's yeast extract, 2% Bacto's peptone, 2% glucose, and 2% agarose. . The agar medium was left to stand at 30 ° C. for 2 days, and a high yield of polyamine was produced among colonies that appeared.

The genome of this high producing strain was extracted with GenTLE (registered trademark) manufactured by Takara Bio Co., Ltd. Subsequently, genome analysis of high-producing strains was performed using Illumina HiSeq, and compared with the arrangement of wild strains, homozygous mutations of four genes related to the synthesis of polyamines were confirmed. That is, the 266th C of the leucine synthetase LEU1 encoded on the third chromosome is A, and the 1964th T of the aconitic acid synthetic gene ACO2 encoded on the 10th chromosome is C, and is encoded on the 12th chromosome. It was found that the 1130th C of the S-adenosyl-L-methionine synthetic gene SAM1 mutated to G, and the 744th A of the aspartic acid / glutamic acid transport gene AGC1 encoded on the 16th chromosome to T. For the leucine synthetic gene LEU1, the aconitate synthetic gene ACO2, the S-adenosyl-L-methionine synthetic gene SAM1, the aspartic acid / glutamic acid transport gene AGC1, the base sequence according to the present invention is shown in SEQ ID NOs: 1, 3, 5, and 7. , Wild type is shown in SEQ ID NOs: 2, 4, 6, 8.

Arginase gene CAR1 coded for chromosome 16, ornithintal carbonate gene SPE1 coded for chromosome 11, spermidine synthetic gene SPE3 coded for chromosome 16, as a polyamine synthetic gene Although the spermine synthetic gene SPE4 encoded on the chromosome is known, no polyamine synthetic gene has been mutated in high producing strains.

(Example 1)

The mutant yeast strain of the polyamine high production was cultured. The culture was 50 g / L sucrose, 3 g / L ammonium sulfate, 4 g / L potassium dihydrogen phosphate, 2 g / L potassium dihydrogen phosphate, 0.4 g / L magnesium sulfate and hexahydrate, 10 g / L bacterium yeast extract, and 0.002 g / biotin. L, calcium chloride, dihydrate 0.4g / L, manganese chloride, tetrahydrate 0.01g / L, zinc sulfate, heptahydrate 0.02g / L, iron citrate, n hydrate 0.06g / L, boric acid 0.002g / L, ammonium molybdate It was carried out in a medium having a composition of 0.001 g / L tetrahydrate, 0.001 g / L potassium iodide, 0.001 g / L copper sulfate pentahydrate, 0.001 g / L cobalt chloride hexahydrate, and 0.05 g / L salt. The mutant yeast was incubated for 4 days with stirring at the culture temperature of 30 ℃. The culture was centrifuged to concentrate the cells and washed again. This was heat-processed at 60 degreeC for 1 hour, and spray-dried. As for the spray dry, the Buze mini spray dryer b-290 was used. Dry yeast was obtained at an outlet temperature of 85-90 ° C. Dry yeast contained 0.72 mg / g putrescine, 3.62 mg / g spermidine and 0.38 mg / g spermine (total 4.72 mg / g).

(Reference example)

S-adenosylmethionine (SAMe) containing yeast was obtained by the method of Unexamined-Japanese-Patent No. 5974891 Paragraph 0014. Putrescine was not detected, and spermidine was 0.29 mg / g and spermine was 0.16 mg / g (total 0.45 mg / g).

(Comparative Example 1)

Dry yeast of Comparative Example 1 was obtained by culturing Saccharomyces (Association 7) under the same conditions as in Example 1. Putrescine was not detected and the content of spermidine was 0.51 mg / g and the content of spermine was 0.45 mg / g (total 0.96 mg / g).

(Example 2)

In order to confirm the effect of the crystalline polysaccharide, in addition to the above examples and comparative examples, a sample to which γ-cyclodextrin was added before the spray dry step was prepared.

Cell cultures were concentrated by centrifugation of a culture yeast obtained in Example 1 (putrescine 0.72 mg / g, spermidine 3.62 mg / g, spermine 0.38 mg / g, total 4.72 mg / g), Washed again. The cells and γ-cyclodextrin were mixed in the medium so that the ratio of the cell mass (bacterial concentration x broth liquid): cyclodextrin weight in the culture was 7: 3. The resulting mixture was treated at 60 ° C. for 1 hour. The polyamine content at this time was 0.55 mg / g of putrescine, 2.63 mg / g of spermidine and 0.29 mg / g of spermine, and the total was 3.47 mg / g. As the cyclodextrin was added, the polyamine content per unit weight was lowered, but the polyamine in the yeast was not removed. Thereafter, the yeast was spray-dried using the Buheze mini spray dryer b-290. Dry yeast was obtained at an outlet temperature of 85-90 ° C. This dry yeast is a powdery composition which contains 30 weight% of cyclodextrins in addition to dry yeast (Hereinafter, dry yeast or its powdery composition is also called yeast powder). The polyamine content was 0.55 mg / g putrescine, 2.63 mg / g spermidine and 0.29 mg / g spermine (total 3.47 mg / g). The polyamine content was not reduced before and after spray drying.

(Example 3)

The mutant yeast of Example 1 was cultured under the same conditions as in Example 1. Centrifuge the culture medium containing the prepared yeast (putrescine 1.07mg / g, spermidine 4.24mg / g, spermine 0.36mg / g, total 5.66mg / g) to concentrate the cells, and washed again It was. An aqueous solution containing α-cyclodextrin was added to the cells so that the α-cyclodextrin was 50% by weight. This was treated at 60 ° C. for 1 hour. The polyamine content at this point was 0.58 mg / g putrescine, 2.18 mg / g spermidine and 0.18 mg / g spermine per dry solid, and the total was 2.93 mg / g. As the cyclodextrin was added, the polyamine content per unit weight was lowered, but the polyamine in the yeast was not removed. The yeast was then spray dried. As for the spray dry, the Buze mini spray dryer b-290 was used. Dry yeast powder was obtained at an outlet temperature of 95 to 100 ° C. The polyamine content in the dry powder was 0.63 mg / g of putrescine, 2.26 mg / g of spermidine and 0.19 mg / g of spermine (total 3.08 mg / g). The polyamine content was not reduced before and after spray drying.

(Example 4)

The yeast culture solution of Example 2 and the yeast culture solution of Comparative Example 1 before cell concentration were mixed in the ratio of 2: 3, and this was processed at 60 degreeC for 1 hour. Thereafter, the yeast culture solution was vacuum dried to obtain dry yeast. As a result, a yeast powder having a content of putrescine of 0.29 mg / g, a content of spermidine of 1.75 mg / g, and a spermine of 0.42 mg / g (total 2.46 mg / g) was obtained.

Cell activation test

2000 Hela cells were seeded in a 96-well plate well, suspended in 100ul of DMEM · 10% fetal bovine serum mixed medium. Next, the yeast powder of Example 1, the yeast powder of Example 2, the yeast powder of Example 4, the yeast powder of Reference Example, the yeast powder of Comparative Example 1, and the soybean extract (Comparative Example 2), Rice germ extract (Comparative Example 3) and yeast extract (Comparative Example 4) were each suspended in water at a concentration of 10000 mg / L and autoclave sterilized. Each liquid was well suspended and 6.3ul was added to the wells. By this operation, the concentration of each sample was 600 mg / L. In addition, 6.3ul of sterilized water was put into another well as a control. 96-well plates were allowed to stand for 20 hours at 37 ° C. under 5% carbon dioxide conditions. Thereafter, the medium was discarded, and each well was again washed with 100ul of DMEM medium. Thereafter, 100 ul of a mixture of DMEM and 8 cell counting kits manufactured by Dojin Chemical Research Institute at a volume ratio of 10: 1 was added to each well, and allowed to stand at 37 ° C for 40 minutes. Finally, 450 nm absorbance of each well was measured using Perkin Elmer ARVO X3. The proliferation of the cells was evaluated by the ratio of absorbance with the control.

TABLE 1

In the high spermidine-containing yeast obtained in the present invention, the cell proliferation activity was particularly high compared to Comparative Examples 2 to 4, and 1.6 to 1.7 times compared to the control. It is thought that the cell activating ability becomes high because a polyamine exists in yeast. Although polyamines have been thought to be the same in vivo in any form, surprisingly, it has been found that the magnitude of the effect varies depending on the type of food provided, and in particular in the form of high polyamine-containing yeast.

150 ℃ heating test

Yeast powder, soybean extract powder (Comparative Example 2), rice germ extract powder (Comparative Example 3) and yeast extract powder (Comparative Example 4) described in Examples 1 and 2 were enclosed in a glass test tube with 20 mg each. After leaving 0 to 60 minutes on the alumina block heater of 150 degreeC, the polyamine content was measured.

TABLE 2

The number at the top is the polyamine content [mg / g] and the number at the bottom is the ratio [%] before heating.

The same heating test was performed on the said yeast powder, and the spermidine content of each powder was measured. The results are shown in Table 3.

TABLE 3

The upper number is the spermidine content [mg / g] and the lower number is the ratio [%] before heating.

The composition (high spermidine-containing yeast) of the present invention was found to be less likely to reduce polyamine even after heating at 150 ° C., compared to conventional polyamine-containing foods. Similarly, the amount of spermidine alone was found to be difficult to reduce. By the present invention, it is thought that polyamine is stabilized by being contained in yeast. The composition (high spermidine-containing yeast) of the present invention is a polyamine food that is difficult to decompose polyamine even when heated at around 150 ° C, and is easily heat-cooked. Moreover, adding cyclodextrin was excellent in heat stability compared with the case of no addition. It is thought that the polyamine in yeast was stabilized by cyclodextrin.

Odor test

To the dry yeast obtained in Example 1, α-cyclodextrin was added so that the weight ratio of yeast: α-cyclodextrin was 1: 1 and mixed as it was in powder (Example 5). The finished food is 50% α-cyclodextrin containing yeast.

1 g of the food was placed in a 250 ml glass container with a lid, sealed, and placed at room temperature for 1 hour, and then covered with a lid to treat yeast in the container. The results are shown in Table 4.

TABLE 4

Yeast odor was improved by using the modified yeast of the present invention in combination with cyclodextrin. Cyclodextrin can improve the stability of a polyamine, increase thermal stability, and can also improve yeast odor along with the effect which makes processing and cooking easy.

Industrial availability

Since the mutant yeast of the present invention contains a large amount of polyamine, in particular spermidine, in cells as compared with the conventional yeast, it is intended to be suitably blended into food and drink intended for cell activation by spermidine. In addition, the modified yeast of the present invention exhibits the effect of not only increasing the thermal stability but also reducing the odor by using together with the crystalline polysaccharide.

SEQUENCE LISTING <110> MITSUBISHI GAS CHEMICAL COMPANY, INC. <120> Yeast with high content of polyamines, and food and drinkcomposition comprising the same <130> 2016-074 <150> JP 2017-053205 <151> 2017-03-17 <160> 8 <170> PatentIn version 3.5 <210> 1 <211> 1095 <212> DNA <213> Saccharomyces cerevisiae <400> 1 atgtctgccc ctaagaagat cgtcgttttg ccaggtgacc acgttggtca agaaatcaca 60 gccgaagcca ttaaggttct taaagctatt tctgatgttc gttccaatgt caagttcgat 120 ttcgaaaatc atttaattgg tggtgctgct atcgatgcta caggtgttcc acttccagat 180 gaggcgctgg aagcctccaa gaaggctgat gccgttttgt taggtgctgt gggtggtcct 240 aaatggggta ccggtagtgt tagacatgaa caaggtttac taaaaatccg taaagaactt 300 caattgtacg ccaacttaag accatgtaac tttgcatccg actctctttt agacttatct 360 ccaatcaagc cacaatttgc taaaggtact gacttcgttg ttgtcagaga attagtggga 420 ggtatttact ttggtaagag aaaggaagac gatggtgatg gtgtcgcttg ggatagtgaa 480 caatacaccg ttccagaagt gcaaagaatc acaagaatgg ccgctttcat ggccctacaa 540 catgagccac cattgcctat ttggtccttg gataaagcta atgttttggc ctcttcaaga 600 ttatggagaa aaactgtgga ggaaaccatc aagaacgaat tccctacatt gaaggttcaa 660 catcaattga ttgattctgc cgccatgatc ctagttaaga acccaaccca cctaaatggt 720 attataatca ccagcaacat gtttggtgat atcatctccg atgaagcctc cgttatccca 780 ggttccttgg gtttgttgcc atctgcgtcc ttggcctctt tgccagacaa gaacaccgca 840 tttggtttgt acgaaccatg ccacggttct gctccagatt tgccaaagaa taaggtcaac 900 cctatcgcca ctatcttgtc tgctgcaatg atgttgaaat tgtcattgaa cttgcctgaa 960 gaaggtaagg ccattgaaga tgcagttaaa aaggttttgg atgcaggtat cagaactggt 1020 gatttaggtg gttccaacag taccaccgaa gtcggtgatg ctgtcgccga agaagttaag 1080 aaaatccttg cttaa 1095 <210> 2 <211> 1095 <212> DNA <213> Saccharomyces cerevisiae <400> 2 atgtctgccc ctaagaagat cgtcgttttg ccaggtgacc acgttggtca agaaatcaca 60 gccgaagcca ttaaggttct taaagctatt tctgatgttc gttccaatgt caagttcgat 120 ttcgaaaatc atttaattgg tggtgctgct atcgatgcta caggtgttcc acttccagat 180 gaggcgctgg aagcctccaa gaaggctgat gccgttttgt taggtgctgt gggtggtcct 240 aaatggggta ccggtagtgt tagacctgaa caaggtttac taaaaatccg taaagaactt 300 caattgtacg ccaacttaag accatgtaac tttgcatccg actctctttt agacttatct 360 ccaatcaagc cacaatttgc taaaggtact gacttcgttg ttgtcagaga attagtggga 420 ggtatttact ttggtaagag aaaggaagac gatggtgatg gtgtcgcttg ggatagtgaa 480 caatacaccg ttccagaagt gcaaagaatc acaagaatgg ccgctttcat ggccctacaa 540 catgagccac cattgcctat ttggtccttg gataaagcta atgttttggc ctcttcaaga 600 ttatggagaa aaactgtgga ggaaaccatc aagaacgaat tccctacatt gaaggttcaa 660 catcaattga ttgattctgc cgccatgatc ctagttaaga acccaaccca cctaaatggt 720 attataatca ccagcaacat gtttggtgat atcatctccg atgaagcctc cgttatccca 780 ggttccttgg gtttgttgcc atctgcgtcc ttggcctctt tgccagacaa gaacaccgca 840 tttggtttgt acgaaccatg ccacggttct gctccagatt tgccaaagaa taaggtcaac 900 cctatcgcca ctatcttgtc tgctgcaatg atgttgaaat tgtcattgaa cttgcctgaa 960 gaaggtaagg ccattgaaga tgcagttaaa aaggttttgg atgcaggtat cagaactggt 1020 gatttaggtg gttccaacag taccaccgaa gtcggtgatg ctgtcgccga agaagttaag 1080 aaaatccttg cttaa 1095 <210> 3 <211> 2370 <212> DNA <213> Saccharomyces cerevisiae <400> 3 atgctatctt cagctaatag gttttatata aagaggcatt tggcaacaca tgccaatatg 60 ttcccctctg tatctaaaaa ttttcaaaca aaagtgccac cttatgcaaa acttttgacc 120 aacctagaca agattaaaca aataacaaac aatgctccat tgacactagc agaaaaaatt 180 ttatactcgc atctttgcga tcctgaagaa tcgattactt cttctgatct gtccaccatc 240 cgtggtaata aatacttgaa gctaaaccca gaccgtgtag ctatgcagga cgcttctgcc 300 caaatggcgc ttctacaatt catgaccacc ggtctaaatc aaacatctgt cccagcatcc 360 atacattgtg atcatttaat tgtaggtaag gacggtgaaa ctaaagattt accctcttcc 420 atagctacca accaagaagt tttcgatttc ttggagagtt gcgcaaagag atatggaatt 480 caattctggg gcccaggttc tggtatcatt caccagattg ttttggaaaa tttctcagct 540 ccaggtctaa tgatgctagg tactgattcc catacaccaa atgcaggcgg tctgggagct 600 attgccatcg gggttggtgg tgcggatgca gttgatgccc tcacaggcac tccatgggaa 660 ttaaaagcac caaaaatttt aggtgttaaa ttgaccggaa agttaaacgg atggtccact 720 cctaaagatg taatcacaaa gctcgctggt ttactaactg tcagaggtgg tactggttat 780 atcgtcgagt acttcggcga aggtgtatcc actctatctt gcacaggtat ggcaaccatc 840 tgtaatatgg gagctgaaat cggtgctaca acgtcaactt tcccttacca agaagctcac 900 aagcgttatt tgcaagcaac taatagagca gaggtcgctg aagcagctga tgttgcttta 960 aacaagttta acttcttaag agccgacaaa gatgctcaat acgataaagt tattgaaatt 1020 gacttatccg caattgaacc tcacgttaat ggtccattta caccagacct ttcaacccca 1080 atatctcaat atgccgaaaa aagtttgaag gaaaactggc cccaaaaagt tagcgctggt 1140 ttgattggat catgtaccaa ttcatcttat caagacatga gtcgtgttgt cgacttggtc 1200 aagcaagctt ccaaagccgg cttgaaacca cgtatcccct tctttgtcac ccctggttca 1260 gaacaaatta gagctacctt ggaaagagat ggaatcatcg atattttcca agaaaatggt 1320 gccaaagttt tagcaaatgc atgcggccct tgtatcggac aatggaatag ggaagatgtc 1380 tcgaaaacat caaaagaaac gaacactatt tttacatcat tcaatagaaa tttcagagct 1440 agaaacgatg gtaataggaa tacaatgaat ttcttaacat ccccagaaat agtaacagcg 1500 atgagttatt ctggagatgc tcagttcaat ccgctaactg actcaattaa attgccaaat 1560 gggaaggatt tcaaattcca accaccaaag ggtgatgagt taccaaaaag aggatttgaa 1620 cacggtagag acaaatttta tcctgaaatg gatccaaagc cagatagcaa tgtagagatt 1680 aaggtagacc ctaattctga tcgtttgcaa ttattagagc cattcaaacc ttggaacggg 1740 aaggaattga agacaaacgt gcttttgaaa gttgaaggta aatgtacaac agatcatatt 1800 tccgctgcgg gcgtctggtt gaaatataaa ggccatctag aaaacatttc ttacaataca 1860 ttgattggtg cacaaaacaa agaaaccggt gaagtcaaca aggcttatga ccttgacgga 1920 actgaatatg atattcctgg tttgatgatg aaacggaaat cagacggtag accatggacc 1980 gtgatagcgg aacataacta tggtgaaggt tccgcaagag agcatgctgc tttgtcacca 2040 agatttttag gcggagagat tcttttagtt aagtcttttg caagaattca tgagacaaac 2100 ttgaagaaac aaggtgtgtt gccattgact tttgccaacg aatctgacta tgataaaata 2160 tcaagcggag atgttttaga aacgttgaac ctagttgaca tgattgctaa ggatggtaat 2220 aacggtggtg aaattgatgt taaaattact aaaccaaacg gtgaatcgtt caccatcaag 2280 gcaaaacata ctatgtctaa agatcaaatc gattttttca aagctggttc agcaatcaat 2340 tatattggta atatacgaag aaacgaataa 2370 <210> 4 <211> 2370 <212> DNA <213> Saccharomyces cerevisiae <400> 4 atgctatctt cagctaatag gttttatata aagaggcatt tggcaacaca tgccaatatg 60 ttcccctctg tatctaaaaa ttttcaaaca aaagtgccac cttatgcaaa acttttgacc 120 aacctagaca agattaaaca aataacaaac aatgctccat tgacactagc agaaaaaatt 180 ttatactcgc atctttgcga tcctgaagaa tcgattactt cttctgatct gtccaccatc 240 cgtggtaata aatacttgaa gctaaaccca gaccgtgtag ctatgcagga cgcttctgcc 300 caaatggcgc ttctacaatt catgaccacc ggtctaaatc aaacatctgt cccagcatcc 360 atacattgtg atcatttaat tgtaggtaag gacggtgaaa ctaaagattt accctcttcc 420 atagctacca accaagaagt tttcgatttc ttggagagtt gcgcaaagag atatggaatt 480 caattctggg gcccaggttc tggtatcatt caccagattg ttttggaaaa tttctcagct 540 ccaggtctaa tgatgctagg tactgattcc catacaccaa atgcaggcgg tctgggagct 600 attgccatcg gggttggtgg tgcggatgca gttgatgccc tcacaggcac tccatgggaa 660 ttaaaagcac caaaaatttt aggtgttaaa ttgaccggaa agttaaacgg atggtccact 720 cctaaagatg taatcacaaa gctcgctggt ttactaactg tcagaggtgg tactggttat 780 atcgtcgagt acttcggcga aggtgtatcc actctatctt gcacaggtat ggcaaccatc 840 tgtaatatgg gagctgaaat cggtgctaca acgtcaactt tcccttacca agaagctcac 900 aagcgttatt tgcaagcaac taatagagca gaggtcgctg aagcagctga tgttgcttta 960 aacaagttta acttcttaag agccgacaaa gatgctcaat acgataaagt tattgaaatt 1020 gacttatccg caattgaacc tcacgttaat ggtccattta caccagacct ttcaacccca 1080 atatctcaat atgccgaaaa aagtttgaag gaaaactggc cccaaaaagt tagcgctggt 1140 ttgattggat catgtaccaa ttcatcttat caagacatga gtcgtgttgt cgacttggtc 1200 aagcaagctt ccaaagccgg cttgaaacca cgtatcccct tctttgtcac ccctggttca 1260 gaacaaatta gagctacctt ggaaagagat ggaatcatcg atattttcca agaaaatggt 1320 gccaaagttt tagcaaatgc atgcggccct tgtatcggac aatggaatag ggaagatgtc 1380 tcgaaaacat caaaagaaac gaacactatt tttacatcat tcaatagaaa tttcagagct 1440 agaaacgatg gtaataggaa tacaatgaat ttcttaacat ccccagaaat agtaacagcg 1500 atgagttatt ctggagatgc tcagttcaat ccgctaactg actcaattaa attgccaaat 1560 gggaaggatt tcaaattcca accaccaaag ggtgatgagt taccaaaaag aggatttgaa 1620 cacggtagag acaaatttta tcctgaaatg gatccaaagc cagatagcaa tgtagagatt 1680 aaggtagacc ctaattctga tcgtttgcaa ttattagagc cattcaaacc ttggaacggg 1740 aaggaattga agacaaacgt gcttttgaaa gttgaaggta aatgtacaac agatcatatt 1800 tccgctgcgg gcgtctggtt gaaatataaa ggccatctag aaaacatttc ttacaataca 1860 ttgattggtg cacaaaacaa agaaaccggt gaagtcaaca aggcttatga ccttgacgga 1920 actgaatatg atattcctgg tttgatgatg aaatggaaat cagacggtag accatggacc 1980 gtgatagcgg aacataacta tggtgaaggt tccgcaagag agcatgctgc tttgtcacca 2040 agatttttag gcggagagat tcttttagtt aagtcttttg caagaattca tgagacaaac 2100 ttgaagaaac aaggtgtgtt gccattgact tttgccaacg aatctgacta tgataaaata 2160 tcaagcggag atgttttaga aacgttgaac ctagttgaca tgattgctaa ggatggtaat 2220 aacggtggtg aaattgatgt taaaattact aaaccaaacg gtgaatcgtt caccatcaag 2280 gcaaaacata ctatgtctaa agatcaaatc gattttttca aagctggttc agcaatcaat 2340 tatattggta atatacgaag aaacgaataa 2370 <210> 5 <211> 1149 <212> DNA <213> Saccharomyces cerevisiae <400> 5 atggccggta catttttatt cacttctgaa tccgttggtg aaggtcaccc agataagatc 60 tgtgaccaag tttccgacgc catcttggac gcttgtttag ccgaggaccc tcactccaaa 120 gttgcgtgtg aaaccgcggc aaagactggt atgattatgg tctttggtga aattactacc 180 aaggcacagt tggattacca aaaaatcgtc agagacacca tcaagaagat tggttacgat 240 gattccgcca agggtttcga ctataagacc tgtaacgtcc ttgtcgccat tgagcaacaa 300 tctccagata tcgcccaagg tgtccacgag gagaaggatt tggaagacat cggtgccggt 360 gaccaaggta tcatgtttgg ttacgccaca gatgaaactc cagagggttt gcctttgact 420 attcttttgg ctcataaact aaacatggcc atggctgacg cgagaagaga tggctcttta 480 gcgtggttga gaccagacac caagactcaa gtcaccgtcg aatacaagga tgaccacggt 540 agatgggttc cacaaagaat cgacaccgtc gtcgtctccg ctcaacatgc tgacgaaatc 600 acgaccgagg acttaagagc gcaactaaag tccgagatca ttgaaaaagt catcccaaga 660 gacatgttgg acgaaaacac caaatacttt atccaacctt ccggtagatt cgtcatcggt 720 ggtcctcaag gtgacgctgg tttgaccggt agaaagatca tcgtcgacgc ttacggtggt 780 gcctcatccg tcggtggtgg tgccttctcc ggtaaggact actctaaggt tgatcgttct 840 gccgcttatg ccgctagatg ggttgccaag tccctagttg ccgctggttt atgtaagaga 900 gttcaagttc aattttctta tgccatcggt attgcggaac cattgtcctt gcacgttgac 960 acctatggta ctgcgaccaa gtctgacgaa gaaattatcg acattatcag caagaacttt 1020 gacttgagac ctggtgtatt ggtcaaggag ttggacttag ctagaccaat ctacttgcca 1080 accgcttctt atggccattt cacaaaccaa gaatacccat gggaaaagcg taagactttg 1140 aagttctaa 1149 <210> 6 <211> 1149 <212> DNA <213> Saccharomyces cerevisiae <400> 6 atggccggta catttttatt cacttctgaa tccgttggtg aaggtcaccc agataagatc 60 tgtgaccaag tttccgacgc catcttggac gcttgtttag ccgaggaccc tcactccaaa 120 gttgcgtgtg aaaccgcggc aaagactggt atgattatgg tctttggtga aattactacc 180 aaggcacagt tggattacca aaaaatcgtc agagacacca tcaagaagat tggttacgat 240 gattccgcca agggtttcga ctataagacc tgtaacgtcc ttgtcgccat tgagcaacaa 300 tctccagata tcgcccaagg tgtccacgag gagaaggatt tggaagacat cggtgccggt 360 gaccaaggta tcatgtttgg ttacgccaca gatgaaactc cagagggttt gcctttgact 420 attcttttgg ctcataaact aaacatggcc atggctgacg cgagaagaga tggctcttta 480 gcgtggttga gaccagacac caagactcaa gtcaccgtcg aatacaagga tgaccacggt 540 agatgggttc cacaaagaat cgacaccgtc gtcgtctccg ctcaacatgc tgacgaaatc 600 acgaccgagg acttaagagc gcaactaaag tccgagatca ttgaaaaagt catcccaaga 660 gacatgttgg acgaaaacac caaatacttt atccaacctt ccggtagatt cgtcatcggt 720 ggtcctcaag gtgacgctgg tttgaccggt agaaagatca tcgtcgacgc ttacggtggt 780 gcctcatccg tcggtggtgg tgccttctcc ggtaaggact actctaaggt tgatcgttct 840 gccgcttatg ccgctagatg ggttgccaag tccctagttg ccgctggttt atgtaagaga 900 gttcaagttc aattttctta tgccatcggt attgcggaac cattgtcctt gcacgttgac 960 acctatggta ctgcgaccaa gtctgacgaa gaaattatcg acattatcag caagaacttt 1020 gacttgagac ctggtgtatt ggtcaaggag ttggacttag ctagaccaat ctacttgcca 1080 accgcttctt atggccattt cacaaaccaa gaatacccat gggaaaagcc taagactttg 1140 aagttctaa 1149 <210> 7 <211> 2709 <212> DNA <213> Saccharomyces cerevisiae <400> 7 atggagcaaa tcaattcgaa cagtagaaaa aagaagcaac aattggaagt attcaaatat 60 tttgcaagtg tccttacaaa agaggacaag cctattagta tcagtaatgg tatgttagat 120 atgccgacag tgaactccag taaactcaca gcaggaaatg ggaaacctga cacggagaag 180 cttacaggag aactaatttt aacatacgac gatttcattg aactgatatc tagctcaaag 240 actatttatt cgaagtttac ggaccattcg ttcaatttga accagatacc caagaacgtt 300 ttcgggtgta ttttcttcgc tattgatgaa caaaacaagg gatatctgac gcttaatgat 360 tggttttatt ttaataattt attagaatat gataattatc atctcattat tctatatgag 420 ttctttagga aatttgatgt agagaatttg aaggcaaaac aaaaaaaaga gcttggtagt 480 tcgtcgttta atttaaaggc tgcagatgat cgaattaagt caattaatta tggtaacaga 540 tttctaagct ttgatgatct tcttttgaat ctgaaccaat tcaaagatac tatccgcctg 600 ttgcacgaat ctattgatga taattttgtt aaagataaca aattactact tgattggaat 660 gactttcgat ttctgaaatt ttacaaatgt tatcatgaaa atgaagagta tttgagttta 720 aactctctgg tcacgatttt acataatgat cttaagaatg aaaaaatatt tataggtttt 780 gataggttgg cacagatgga ctcacaaggg catcgtttag ccctaagcaa aaatcaactc 840 acctatcttc taaggttatt ttactctcac agggtgtctg cagatatatt ttcctccttg 900 aatctatcaa acaccgaatt actaaaagcg gacaataatt ccattccgta caatgtattc 960 aaggatatat tttatttatt tcaaaatttt gacctactga accaaatatt tcacaagtat 1020 gttactgaaa ataatttgaa tgagcaggat attagggaac aaatagttac taaaaatgac 1080 tttatgacag ttttaaacgc ccagtataac aaagtaaaca atatcattga gttctctcct 1140 tcccaaatca acctactatt ttctatcgtc gcaaattcaa aggaaaacag aagattaaga 1200 aagagaaatc aagatcgaga tgacgagcta ttaaatgatc accattatga ttcagatatt 1260 gattttttta tccataatga gtatttgcat ggagtaagca gatccagaaa aaatttagaa 1320 agttttaatg actattatca tgatctctcg gatggatttg accaagactc tggtgttaaa 1380 aaagcttcaa aagcgagtac tggcttgttt gaatctgtat ttggaggtaa aaaagataaa 1440 gcaacgatgc gttctgactt aacaattgaa gatttcatga aaattttgaa cccaaattac 1500 ctgaacgact tagttcacca aatggaattg caaaaaaatc aaaatgagtc attgtatatt 1560 aattactact tttatccaat tttcgattcg ttgtacaatt tctccttggg ttctattgcg 1620 ggttgtattg gtgcaactgt agtataccca atagacttta taaaaacaag gatgcaagcc 1680 caaagatctt tagcccaata caaaaactca attgattgtt tgttgaagat tatatcccgc 1740 gaaggaataa aaggtctcta ctctggctta gggccacaat taataggagt tgctcctgaa 1800 aaggcgataa aattgactgt caatgatttt atgagaaaca ggttgactga taaaaacggc 1860 aagctaagcc tttttcctga aattatttct ggcgcttcag ctggtgcatg tcaagttata 1920 tttactaatc cgttagagat tgtaaaaatt aggctacagg tccaatccga ctatgttggt 1980 gaaaacatac aacaagccaa tgaaactgcc actcaaatag tcaaaaaatt aggactgagg 2040 ggcttgtaca atggtgtagc cgcatgttta atgagagatg ttccattctc tgctatttat 2100 tttcccactt atgcacattt aaaaaaagat ctctttgatt ttgatccaaa tgataaaaca 2160 aagaggaatc gattaaaaac atgggagctt ttaactgccg gtgccattgc tggtatgcca 2220 gctgccttct tgactactcc ttttgatgtt ataaaaacaa ggctccagat agatcctcga 2280 aaaggtgaga caaagtataa cggtatattt catgctatcc gaactatctt aaaggaagag 2340 agctttagaa gctttttcaa aggtggtgga gcccgtgtcc taagaagttc tccccaattt 2400 gggttcactc tggccgccta tgaattattc aagggcttta ttccctcccc cgataacaaa 2460 ttaaaaagca gagagggtag gaagagattt tgtatcgatg acgacgcagg caatgaagag 2520 acagtagttc atagtaacgg tgaactccca cagcaaaagt tttactctga tgatagaaaa 2580 catgccaatt attactataa aagctgtcaa attgcgaaaa cattcattga tttggacaat 2640 aacttttcta ggtttgactc ttcagtttat aaaaactttc aagagcacct aagaagcatt 2700 aacgggtga 2709 <210> 8 <211> 2709 <212> DNA <213> Saccharomyces cerevisiae <400> 8 atggagcaaa tcaattcgaa cagtagaaaa aagaagcaac aattggaagt attcaaatat 60 tttgcaagtg tccttacaaa agaggacaag cctattagta tcagtaatgg tatgttagat 120 atgccgacag tgaactccag taaactcaca gcaggaaatg ggaaacctga cacggagaag 180 cttacaggag aactaatttt aacatacgac gatttcattg aactgatatc tagctcaaag 240 actatttatt cgaagtttac ggaccattcg ttcaatttga accagatacc caagaacgtt 300 ttcgggtgta ttttcttcgc tattgatgaa caaaacaagg gatatctgac gcttaatgat 360 tggttttatt ttaataattt attagaatat gataattatc atctcattat tctatatgag 420 ttctttagga aatttgatgt agagaatttg aaggcaaaac aaaaaaaaga gcttggtagt 480 tcgtcgttta atttaaaggc tgcagatgat cgaattaagt caattaatta tggtaacaga 540 tttctaagct ttgatgatct tcttttgaat ctgaaccaat tcaaagatac tatccgcctg 600 ttgcacgaat ctattgatga taattttgtt aaagataaca aattactact tgattggaat 660 gactttcgat ttctgaaatt ttacaaatgt tatcatgaaa atgaagagta tttgagttta 720 aactctctgg tcacgatttt acaaaatgat cttaagaatg aaaaaatatt tataggtttt 780 gataggttgg cacagatgga ctcacaaggg catcgtttag ccctaagcaa aaatcaactc 840 acctatcttc taaggttatt ttactctcac agggtgtctg cagatatatt ttcctccttg 900 aatctatcaa acaccgaatt actaaaagcg gacaataatt ccattccgta caatgtattc 960 aaggatatat tttatttatt tcaaaatttt gacctactga accaaatatt tcacaagtat 1020 gttactgaaa ataatttgaa tgagcaggat attagggaac aaatagttac taaaaatgac 1080 tttatgacag ttttaaacgc ccagtataac aaagtaaaca atatcattga gttctctcct 1140 tcccaaatca acctactatt ttctatcgtc gcaaattcaa aggaaaacag aagattaaga 1200 aagagaaatc aagatcgaga tgacgagcta ttaaatgatc accattatga ttcagatatt 1260 gattttttta tccataatga gtatttgcat ggagtaagca gatccagaaa aaatttagaa 1320 agttttaatg actattatca tgatctctcg gatggatttg accaagactc tggtgttaaa 1380 aaagcttcaa aagcgagtac tggcttgttt gaatctgtat ttggaggtaa aaaagataaa 1440 gcaacgatgc gttctgactt aacaattgaa gatttcatga aaattttgaa cccaaattac 1500 ctgaacgact tagttcacca aatggaattg caaaaaaatc aaaatgagtc attgtatatt 1560 aattactact tttatccaat tttcgattcg ttgtacaatt tctccttggg ttctattgcg 1620 ggttgtattg gtgcaactgt agtataccca atagacttta taaaaacaag gatgcaagcc 1680 caaagatctt tagcccaata caaaaactca attgattgtt tgttgaagat tatatcccgc 1740 gaaggaataa aaggtctcta ctctggctta gggccacaat taataggagt tgctcctgaa 1800 aaggcgataa aattgactgt caatgatttt atgagaaaca ggttgactga taaaaacggc 1860 aagctaagcc tttttcctga aattatttct ggcgcttcag ctggtgcatg tcaagttata 1920 tttactaatc cgttagagat tgtaaaaatt aggctacagg tccaatccga ctatgttggt 1980 gaaaacatac aacaagccaa tgaaactgcc actcaaatag tcaaaaaatt aggactgagg 2040 ggcttgtaca atggtgtagc cgcatgttta atgagagatg ttccattctc tgctatttat 2100 tttcccactt atgcacattt aaaaaaagat ctctttgatt ttgatccaaa tgataaaaca 2160 aagaggaatc gattaaaaac atgggagctt ttaactgccg gtgccattgc tggtatgcca 2220 gctgccttct tgactactcc ttttgatgtt ataaaaacaa ggctccagat agatcctcga 2280 aaaggtgaga caaagtataa cggtatattt catgctatcc gaactatctt aaaggaagag 2340 agctttagaa gctttttcaa aggtggtgga gcccgtgtcc taagaagttc tccccaattt 2400 gggttcactc tggccgccta tgaattattc aagggcttta ttccctcccc cgataacaaa 2460 ttaaaaagca gagagggtag gaagagattt tgtatcgatg acgacgcagg caatgaagag 2520 acagtagttc atagtaacgg tgaactccca cagcaaaagt tttactctga tgatagaaaa 2580 catgccaatt attactataa aagctgtcaa attgcgaaaa cattcattga tttggacaat 2640 aacttttcta ggtttgactc ttcagtttat aaaaactttc aagagcacct aagaagcatt 2700 aacgggtga 2709

Claims (9)

In comparison with Association No. 7 yeast,
In any one or more amino acid sequence of leucine synthase, aconitate synthase, S-adenosyl-L-methionine synthase, or aspartic acid / glutamic acid transport protein, one or several amino acids are substituted, deleted, inserted Or in addition, or in combination of two or more of these, or
One or more nucleotide sequences of the LEU1 gene encoded on the third chromosome, the ACO2 gene encoded on the chromosome 10, the SAM1 gene encoded on the chromosome 12, and the AGC1 gene encoded on the chromosome 16 Wherein one or several bases are substituted, deleted, inserted or added, or mutated to two or more of these,
Furthermore, yeast variant which has high polyamine.
The method of claim 1,
Any one of the bases at the positions corresponding to the 266th C of the LEU1 gene, the 1964th T of the ACO2 gene, the 1130th C of the SAM1 gene, and the 744th A of the AGC1 gene in association 7 yeast Or yeast variants in which more than one base is substituted.
The method of claim 2,
The base corresponding to the 266th C of the LEU1 gene is A, and the base corresponding to the 1964th T of the ACO2 gene, which is encoded on the 10th chromosome, is C, and the 1130th C of the SAM1 gene, which is encoded on the 12th chromosome. The yeast variant whose base corresponds to G, the 744th A of AGC1 gene encoded by the 16th chromosome, is T.
The method according to any one of claims 1 to 3,
Yeast variant, which is a yeast belonging to Saccharomyces.
The method according to any one of claims 1 to 4,
Yeast variant whose content of polyamine is a total amount of putrescine, spermidine and spermine.
Food and drink composition containing the yeast variant of any one of Claims 1-5, and a crystalline polysaccharide.
The method of claim 6,
The composition wherein the weight ratio of the yeast variant to the crystalline polysaccharide is 1: 0.1 to 1.
The method according to claim 6 or 7,
Wherein the crystalline polysaccharide is cyclodextrin and / or indigestible dextrin.
In comparison with Association No. 7 yeast,
In any one or more amino acid sequence of leucine synthase, aconitate synthase, S-adenosyl-L-methionine synthase, or aspartic acid / glutamic acid transport protein, one or several amino acids are substituted, deleted, inserted Or in addition, or in combination with two or more of these, or
One or more nucleotide sequences of the LEU1 gene encoded on the third chromosome, the ACO2 gene encoded on the chromosome 10, the SAM1 gene encoded on the chromosome 12, and the AGC1 gene encoded on the chromosome 16 Wherein one or several bases are substituted, deleted, inserted or added, or mutated to a combination of two or more thereof,
A method for producing a yeast variant having high polyamine.

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