EP-4741410-A1 - SOLUBILIZATION GROUP FOR POORLY WATER-SOLUBLE PROTEINS

Abstract

The object of the present invention is to provide novel means that can be used for solubilizing proteins with poor water solubility. Provided is a hydrophilic solubilizing group for proteins or peptides, comprising a sugar chain moiety; and a functional group moiety that is bound to the amino or carboxy group of said sugar chain, and that binds to the amino acid side chain and can be detached from said protein or amino acid side chain in the presence of an acid.

Inventors

• KAJIHARA, YASUHIRO
• OKAMOTO, RYO
• MAKI, Yuta
• LIU, Yanbo
• FUJITA, SHOICHI
• ELOUALI, Sofia

Assignees

• Glytech, Inc.

Dates

Publication Date

20260513

Application Date

20240704

Claims (18)

1. A hydrophilic solubilizing group for proteins or peptides, comprising a sugar chain moiety; and a functional group moiety that is bound to the amino or carboxy group of said sugar chain, and that binds to the amino acid side chain and can be detached from said protein or amino acid side chain in the presence of an acid.
2. The solubilizing group according to Claim 1, comprising a sugar chain moiety; and a triphenylmethanol moiety bound to the amino or carboxy group of said sugar chain.
3. The solubilizing group according to Claim 2, wherein said triphenylmethanol moiety is bound to the carboxy group of said sugar chain.
4. The solubilizing group according to Claim 1, wherein said sugar chain is characterized in that it consists of 4 or more sugars.
5. The solubilizing group according to Claim 1, wherein said sugar chain is characterized in that it consists of 7 or more sugars.
6. The solubilizing group according to Claim 1, wherein said sugar chain is characterized in that it consists of 9 or more sugars.
7. The solubilizing group according to Claim 1, wherein said sugar chain is characterized in that it is a complex type bianntennary sugar chain, a complex type triantennary sugar chain, or a complex type tetraantennary sugar chain.
8. The solubilizing group according to Claim 1, wherein said sugar chain is characterized in that it is a complex type bianntennary sugar chain.
9. The solubilizing group according to Claim 8, wherein said complex type bianntennary sugar chain is characterized in that it is a sugar chain selected from the group consisting of a disialo sugar chain, a monosialo sugar chain, an asialo sugar chain, a diGlcNAc sugar chain, and a dimannose sugar chain.
10. The solubilizing group according to Claim 1, wherein said sugar chain is characterized in that it is a sugar chain represented by the following formula: [wherein R 1 and R 2 are the same or different, to indicate and Ac indicates an acetyl group].
11. The solubilizing group according to Claim 1, having the following structure.
12. A peptide or protein having one or more of the solubilizing groups according to Claim 1.
13. The peptide or protein according to Claim 12, which has poor water solubility.
14. The peptide or protein according to Claim 12, selected from the group consisting of interleukin and transmembrane protein, and a fragment thereof.
15. The peptide or protein according to Claim 14, which is rhodopsin or IL-6, or a fragment thereof.
16. A method for producing a solubilized protein or peptide, comprising a step of binding the solubilizing group according to Claim 1 to the sulfhydryl group, the carboxy group, or the amino group of one or more amino acid side chains constituting any protein or peptide.
17. A method for producing the protein or peptide of interest employing proteins or peptides with poor water solubility, comprising: a step of binding the solubilizing group according to Claim 1 to the sulfhydryl group, the carboxy group, or the amino group of one or more amino acid side chains constituting said proteins or peptides with poor water solubility to allow solubilization, and a step of linking said solubilized protein or peptide with another soluble protein or peptide, or solubilized protein or peptide, to obtain the protein of interest.
18. The method according to claim 17, further comprising: a step of detaching said solubilizing group from said protein of interest in the presence of an acid, and a step of collecting said solubilizing group.

Description

Technical Field The present disclosure relates to a solubilizing group for solubilizing proteins with poor water solubility, as well as a method for producing solubilized proteins etc. employing the said solubilizing group. Background Art There has conventionally been great need for solubilization of proteins. For example, when a protein is to be used as a pharmaceutical active ingredient, it is preferred that the protein is sufficiently soluble in water, in terms of e.g. improvement in absorbability by systemic administration (such as oral administration), enrichment of the active ingredient in formulation, and realization of appropriate viscosity in formulation. Moreover, even at the drug discovery stage, solubilization of proteins is again necessary for protein folding or analysis of complexes of folded proteins and disease-causing target molecules. On the other hand, due to advancement and sophistication of recent drug discovery technology, candidate molecules with poor water solubility are increasingly growing, and the need for solubilization technology is further being escalated. As an approach for solubilizing proteins with poor water solubility, an approach employing a hydrophilic tag is known. For example, Patent Literature 1 describes a hydrophilic polylysine tag for solubilizing proteins with poor water solubility. Citation List Non-Patent Literature [Non-Patent Literature 1] Tsuda et al., Angew. Chem. Int. Ed. 2018, 57, 2105-2109 Summary of the Invention Problems to be Solved by the Invention The object of the present disclosure is to provide novel means that can be used for solubilizing proteins with poor water solubility. Means for Solving the Problems Sugar chains are known to bind to membrane proteins or secretory proteins etc. in vivo, to have various influences on their biological activity. On the other hand, sugar chains, in contrast to their usefulness, were restricted in their use due to difficulty in acquisition and high cost. The inventors of the present application, in the process of repeated extensive investigation in order to solve the above problem, found that proteins with poor water solubility can be solubilized by adding a sugar chain solubilizing group to proteins with poor water solubility. Further, in doing so, it was found that by retaining a particular functional group moiety on the sugar chain solubilizing group, the sugar chain solubilizing group employed for solubilization can be separated/collected and re-utilized after obtaining the solubilized protein of interest, thus coming to complete the invention. That is, the present disclosure encompasses the following characteristics: (1) A hydrophilic solubilizing group for proteins or peptides, comprising a sugar chain moiety; and a functional group moiety that is bound to the amino or carboxy group of said sugar chain, and that binds to the amino acid side chain and can be detached from said protein or amino acid side chain in the presence of an acid.(2) The solubilizing group according to (1), comprising a sugar chain moiety; and a triphenylmethanol moiety bound to the amino or carboxy group of said sugar chain.(3) The solubilizing group according to (2), wherein said triphenylmethanol moiety is bound to the carboxy group of said sugar chain.(4) The solubilizing group according to (1), wherein said sugar chain is characterized in that it consists of 4 or more sugars.(5) The solubilizing group according to (1), wherein said sugar chain is characterized in that it consists of 7 or more sugars.(6) The solubilizing group according to (1), wherein said sugar chain is characterized in that it consists of 9 or more sugars.(7) The solubilizing group according to (1), wherein said sugar chain is characterized in that it is a complex type bianntennary sugar chain, a complex type triantennary sugar chain, or a complex type tetraantennary sugar chain.(8) The solubilizing group according to (1), wherein said sugar chain is characterized in that it is a complex type bianntennary sugar chain.(9) The solubilizing group according to (8), wherein said complex type bianntennary sugar chain is characterized in that it is a sugar chain selected from the group consisting of a disialo sugar chain, a monosialo sugar chain, an asialo sugar chain, a diGlcNAc sugar chain, and a dimannose sugar chain.(10) The solubilizing group according to (1), wherein said sugar chain is characterized in that it is a sugar chain represented by the following formula: [wherein R1 and R2 are the same or different, to indicate , and Ac indicates an acetyl group].(11) The solubilizing group according to (1), having the following structure. (12) A peptide or protein having one or more of the solubilizing groups according to (1).(13) The peptide or protein according to (12), which has poor water solubility.(14) The peptide or protein according to (12), selected from the group consisting of interleukin and transmembrane protein, and as a fragment thereof.(15) The pepti