Gene Encoding Cell Wall Mannoprotein and use Thererof
The present invention relates to a brewer's yeast which produces alcoholic beverages having an ability for reducing the haze level, alcoholic beverages produced using such a yeast, and a method of producing such alcoholic beverages. More specifically, the present invention relates to a yeast which can reduce the level of haze in the product by increasing the level of expression of ScCWP2 gene encoding cell wall mannoprotein Cwp2p in brewer's yeast, or non-ScCWP2 gene characteristic to beer yeast, and to a method of producing alcoholic beverages using such a yeast.
1 . A polynucleotide selected from the group consisting of:
(a) a polynucleotide comprising a polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 1;
(b) a polynucleotide comprising a polynucleotide encoding a protein consisting of the amino acid sequence of SEQ ID NO: 2;
(c) a polynucleotide comprising a polynucleotide encoding a protein consisting of the amino acid sequence of SEQ ID NO: 2 with one or more amino acids thereof being deleted, substituted, inserted and/or added, and functioning as a cell wall mannoprotein;
(d) a polynucleotide comprising a polynucleotide encoding a protein having an amino acid sequence having 60% or higher identity with the amino acid sequence of SEQ ID NO: 2, and functioning as a cell wall mannoprotein;
(e) a polynucleotide comprising a polynucleotide which hybridizes to a polynucleotide consisting of a nucleotide sequence complementary to the nucleotide sequence of SEQ ID NO: 1 under stringent conditions, and which encodes a protein functioning as a cell wall mannoprotein; and
(f) a polynucleotide comprising a polynucleotide which hybridizes to a polynucleotide consisting of a nucleotide sequence complementary to the nucleotide sequence of the polynucleotide encoding the protein of the amino acid sequence of SEQ ID NO: 2 under stringent conditions, and which encodes a protein functioning as a cell wall mannoprotein.
2 . The polynucleotide of claim 1 selected from the group consisting of:
(g) a polynucleotide comprising a polynucleotide encoding a protein consisting of the amino acid sequence of SEQ ID NO: 2, or encoding an amino acid sequence of SEQ ID NO: 2 wherein 1 to 10 amino acids thereof is deleted, substituted, inserted, and/or added, and wherein said protein functions as a cell wall mannoprotein;
(h) a polynucleotide comprising a polynucleotide encoding a protein having 90% or higher identity with the amino acid sequence of SEQ ID NO: 2, and functioning as a cell wall mannoprotein; and
(i) a polynucleotide comprising a polynucleotide which hybridizes to SEQ ID NO: 1 or which hybridizes to a nucleotide sequence complementary to the nucleotide sequence of SEQ ID NO: 1 under stringent conditions, and which encodes a protein functioning as a cell wall mannoprotein.
3 . The polynucleotide of claim 1 comprising a polynucleotide consisting of SEQ ID NO: 1.
4 . The polynucleotide of claim 1 comprising a polynucleotide encoding a protein consisting of SEQ ID NO: 2.
5 . The polynucleotide of claim 1 , wherein the polynucleotide is DNA.
6 . A protein encoded by the polynucleotide of claim 1 .
7 . A vector comprising the polynucleotide of claim 1 .
8 . A vector comprising the polynucleotide selected from the group consisting of:
(j) a polynucleotide comprising a polynucleotide encoding a protein consisting of the amino acid sequence of SEQ ID NO: 4, or encoding an amino acid sequence of SEQ ID NO: 4 wherein 1 to 10 amino acids thereof is deleted, substituted, inserted, and/or added, and wherein said protein functions as a cell wall mannoprotein;
(k) a polynucleotide comprising a polynucleotide encoding a protein having 90% or higher identity with the amino acid sequence of SEQ ID NO: 4, and functioning as a cell wall mannoprotein; and
(l) a polynucleotide comprising a polynucleotide which hybridizes to SEQ ID NO: 3 or which hybridizes to a nucleotide sequence complementary to the nucleotide sequence of SEQ ID NO: 3 under high stringent conditions, and which encodes a protein functioning as a cell wall mannoprotein.
9 . A yeast comprising the vector of claim 7 .
10 . The yeast of claim 9 , wherein a haze-producing ability is reduced by introducing the vector of comprising the polynucleotide.
11 . The yeast of claim 10 , wherein a haze-producing ability is reduced by increasing an expression level of the protein encoded by the polynucleotide.
12 . A method for producing an alcoholic beverage comprising culturing the yeast of claim 9 .
13 . The method for producing an alcoholic beverage of claim 12 , wherein the brewed alcoholic beverage is a malt beverage.
14 . The method for producing an alcoholic beverage of claim 12 , wherein the brewed alcoholic beverage is wine.
15 . An alcoholic beverage produced by the method of claim 12 .
16 . A method for assessing a test yeast for its haze-producing capability, comprising using a primer or a probe designed based on a nucleotide sequence of a gene encoding a cell wall mannoprotein having the nucleotide sequence of SEQ ID NO: 1 or SEQ ID NO: 3.
17 . A method for assessing a test yeast for its haze-producing capability, comprising: culturing a test yeast; and measuring an expression level of a gene encoding a cell wall mannoprotein having the nucleotide sequence of SEQ ID NO: 1 or SEQ ID NO: 3.
18 . A method for selecting a yeast, comprising: culturing test yeasts; quantifying the protein according to Claim 6 or measuring an expression level of a gene encoding a cell wall mannoprotein having the nucleotide sequence of SEQ ID NO: 1 or SEQ ID NO: 3; and selecting a test yeast having said protein amount or said gene expression level according to a target haze-producing capability.
19 . The method for selecting a yeast according to claim 18 , comprising: culturing a reference yeast and test yeasts; measuring an expression level of a gene encoding a cell wall mannoprotein having the nucleotide sequence of SEQ ID NO: 1 or SEQ ID NO: 3 in each yeast; and selecting a test yeast having the gene expressed higher than that in the reference yeast.
20 . The method for selecting a yeast according to claim 18 , comprising: culturing a reference yeast and test yeasts; quantifying the protein encoded by the polynucleotide in each yeast; and selecting a test yeast having said protein for a larger amount than that in the reference yeast.
21 . A method for producing an alcoholic beverage comprising:
conducting fermentation for producing an alcoholic beverage using the yeast according to claim 9 or a yeast selected by the method according to claim 18 ; and adjusting the production amount of haze.