EXTRACTION METHOD FOR HYDROPHILIC RECOMBINANT PROTEIN
The present invention relates to a method for extracting a hydrophilic recombinant protein from a host expressing a hydrophilic recombinant protein. The method includes: a lysate acquisition step in which a dissolving solvent is added to the host expressing a hydrophilic recombinant protein so as to obtain a host cell lysate; and a step in which a diluting solvent is added to the host cell lysate obtained in the lysate acquisition step, an insoluble substance is separated from the obtained dilution, and a supernatant containing the hydrophilic recombinant protein is collected. The hydrophilic recombinant protein has a hydropathy index of 0 or less. The dissolving solvent is an aprotic polar solvent. The diluting solvent is an aqueous solvent.
1 . A method for extracting a hydrophilic recombinant protein from a host expressing a hydrophilic recombinant protein,
the method comprising:
a lysate acquisition step in which a dissolving solvent is added to the host expressing a hydrophilic recombinant protein so as to obtain a host cell lysate; and
a step in which a diluting solvent is added to the host cell lysate obtained in the lysate acquisition step, an insoluble substance is separated from the obtained dilution, and a supernatant containing the hydrophilic recombinant protein is collected,
wherein the hydrophilic recombinant protein has a hydropathy index of 0 or less,
the dissolving solvent is an aprotic polar solvent, and
the diluting solvent is an aqueous solvent.
2 . The method for extracting a hydrophilic recombinant protein according to claim 1 , wherein the dissolving solvent is an aprotic polar solvent not having a cyclic structure.
3 . The method for extracting a hydrophilic recombinant protein according to claim 1 , wherein the dissolving solvent has a dipole moment of 3.0 D or more.
4 . The method for extracting a hydrophilic recombinant protein according to claim 1 , wherein the dissolving solvent is one or more kinds of aprotic polar solvent selected from the group consisting of dimethyl sulfoxide, N,N-dimethylformamide, and N,N-dimethylacetamide.
5 . The method for extracting a hydrophilic recombinant protein according to claim 1 , wherein the dissolving solvent is a solvent obtained by adding an inorganic salt to the aprotic polar solvent.
6 . The method for extracting a hydrophilic recombinant protein according to claim 1 , wherein the diluting solvent is one kind of aqueous solvent selected from the group consisting of (1) water, (2) a mixture of water and ethanol, and (3) a water-soluble buffer solution.
7 . The method for extracting a hydrophilic recombinant protein according to claim 1 , wherein an addition amount of the dissolving solvent is 0.5 times or more a mass of the host by volume (mL)/mass (g) ratio.
8 . The method for extracting a hydrophilic recombinant protein according to claim 1 , wherein an addition amount of the diluting solvent is 0.5 times or more an amount of the host cell lysate by volume ratio.
9 . The method for extracting a hydrophilic recombinant protein according to claim 1 , wherein the hydrophilic protein is a protein selected from the group consisting of a recombinant spider silk protein, a recombinant resilin, and a recombinant collagen.
10 . The method for extracting a hydrophilic recombinant protein according to claim 2 , wherein the dissolving solvent has a dipole moment of 3.0 D or more.
11 . The method for extracting a hydrophilic recombinant protein according to claim 2 , wherein the dissolving solvent is one or more kinds of aprotic polar solvent selected from the group consisting of dimethyl sulfoxide, N,N-dimethylformamide, and N,N-dimethylacetamide.
12 . The method for extracting a hydrophilic recombinant protein according to claim 2 , wherein the dissolving solvent is a solvent obtained by adding an inorganic salt to the aprotic polar solvent.
13 . The method for extracting a hydrophilic recombinant protein according to claim 3 , wherein the dissolving solvent is a solvent obtained by adding an inorganic salt to the aprotic polar solvent.
14 . The method for extracting a hydrophilic recombinant protein according to claim 11 , wherein the dissolving solvent is a solvent obtained by adding an inorganic salt to the aprotic polar solvent.
15 . The method for extracting a hydrophilic recombinant protein according to claim 2 , wherein the diluting solvent is one kind of aqueous solvent selected from the group consisting of (1) water, (2) a mixture of water and ethanol, and (3) a water-soluble buffer solution.
16 . The method for extracting a hydrophilic recombinant protein according to claim 3 , wherein the diluting solvent is one kind of aqueous solvent selected from the group consisting of (1) water, (2) a mixture of water and ethanol, and (3) a water-soluble buffer solution.
17 . The method for extracting a hydrophilic recombinant protein according to claim 11 , wherein the diluting solvent is one kind of aqueous solvent selected from the group consisting of (1) water, (2) a mixture of water and ethanol, and (3) a water-soluble buffer solution.
18 . The method for extracting a hydrophilic recombinant protein according to claim 2 , wherein an addition amount of the dissolving solvent is 0.5 times or more a mass of the host by volume (mL)/mass (g) ratio.
19 . The method for extracting a hydrophilic recombinant protein according to claim 2 , wherein an addition amount of the diluting solvent is 0.5 times or more an amount of the host cell lysate by volume ratio.
20 . The method for extracting a hydrophilic recombinant protein according to claim 2 , wherein the hydrophilic protein is a protein selected from the group consisting of a recombinant spider silk protein, a recombinant resilin, and a recombinant collagen.