TECHNICAL FIELD

The present invention relates to a method for manufacturing an amino acid polymer, in particular to a method for manufacturing a polymer of amino acids linked by peptide bonds with thioacid amino acids.

BACKGROUND ART

Amino acid polymers are utilized for various purposes such as many industrial products, additives to these products, as well as synthetic intermediates of compounds contained in these products. For example, polyaspartic acid which is a homopolymer of aspartic acid is used as detergents, fertilizers, agrochemicals, cosmetics, or additives to these products. Moreover, in recent years, polyalanine contained in spider's thread has gathered attention in regards to its strength. In this way, various use has been found for amino acid polymers as functional high molecular materials that render new characteristics which were not conventionally possessed, and it is anticipated that there will be increasing needs for industrially manufacturing amino acid polymers in years to come.

Various methods such as biological and chemical approaches have been developed as methods for manufacturing amino acid polymers, although room for improvement is recognized in various points such as manufacturing efficiency and quality.

For example, for biological approach, this includes e.g. the restriction for introducing non-natural type amino acids, or the ununiformity of sugar chain structure in glycoproteins.

Moreover, for chemical approach, as opposed to biological approach, freedom of design and structural control are allowed, but steps of protection and deprotection during synthesis are required. For this reason, side reactions that occur during those steps, as well as restriction to productivity such as cost and time are pointed out as problems. It is also pointed out as a problem that the use of toxins with high toxicity is often times necessary in the deprotection step.

Accordingly, in order to improve the productivity of chemical synthesis that allows freedom of design and structural control, it is desired to improve the protection and deprotection steps to simpler and more rapid as well as safer steps.

Thioacid is a generic term for compounds having a thioacid group (—COSH). Methods for forming amide bonds with thioacid thus far known are coupling with an amino acid having an azide group (Non-Patent Literature 1), a method of utilizing thioacid S—S dimer as active ester (Non-Patent Literature 2), and a method of activating by nitrosation of thioacid (Non-Patent Literature 3).

Moreover, methods known for forming peptide bonds using thioacid amino acid are a method of using protected thioacid amino acid S—S dimer as active ester (Non-Patent Literature 4), and a method of utilizing thioacid amino acid as HOBt ester precursor (Non-Patent Literature 5).

CITATION LIST

[Non-Patent Literature 1] J. Am. Chem. Soc. 2003, 125, 7754-7755[Non-Patent Literature 2] Nature, Vol 389, 4 Sep. 1997[Non-Patent Literature 3] Org. Lett., Vol. 13, No. 5, 2011[Non-Patent Literature 4] J. Org. Chem. 2014, 79, 2377-2383[Non-Patent Literature 5] J. Am. Chem. Soc. 9Vol. 132, No. 47, 2010

SUMMARY OF THE INVENTION

Problems to be Solved by the Invention

The object of the present invention is to provide a method for manufacturing an amino acid polymer more simply and efficiently compared to conventional methods for manufacturing amino acid polymers.

Means for Solving the Problems

Peptide bonds between amino acids are formed by using the α-amino group of one amino acid as the nucleophilic group to allow condensation with the carboxyl group of the other amino acid. Since the carboxyl group activated during condensation will become more likely to react with α-amino groups as well as various functional groups having nucleophilicity present in the side chain, it is necessary to protect the side chains and α-amino groups (those other than the subject of reaction) of amino acids in order to manufacture a polymer of structurally controlled amino acids.

As a result of repeated extensive investigation by the present inventors in order to solve the problems of the present invention, it was surprisingly found that an amino acid polymer linked by peptide bonds can be efficiently manufactured without protecting α-amino groups and side chains when thioacid amino acid is employed for the manufacturing of an amino acid polymer and subjecting them to an oxidation reaction with each other. The present inventors further surprisingly found that an amino acid polymer can be similarly manufactured even when using lysine which possess a functional group having a long side chain and nucleophilicity and thus thought that protection was requisite in conventional amino acid condensation methods. This means that the manufacturing method of the present invention can be used for various purposes without being restricted by the type of amino acid side chain.

The present invention is based on the above knowledge, and encompasses the following characteristics:

[1] A method for manufacturing a polymer of amino acids linked by peptide bonds, characterized in that it comprises:

(A) a step of preparing first and second amino acid derivatives each shown below:
H2N—CH(R)—COSH  (1)
(wherein R represents the side chain of any amino acid); and

(B) a step of subjecting said first amino acid derivative and second amino acid derivative to an oxidation reaction to obtain the compound shown below:
H2N—CH(R)—CO—HN—CH(R)—CO—SH  (2)
(wherein each R, each independently, represents the side chain of any amino acid);

where at least one of said first amino acid derivative and second amino acid derivative does not possess a protecting group.

[2] The method according to manufacturing method [1], wherein said first amino acid derivative and second amino acid derivative both do not possess a protecting group.

[3] The manufacturing method according to [1] or [2], comprising:

(C) a step of subjecting Compound (2) obtained in step (B) to an oxidation reaction with a third amino acid derivative shown below:
H2N—CH(R)—COSH  (1)
(wherein R represents the side chain of any amino acid) to obtain the compound shown below:
H2N—CH(R)—CO—(HN—CH(R)—CO)2—SH  (3)
(wherein each R, each independently, represents the side chain of any amino acid).
[4] The manufacturing method according to [3], characterized in carrying out step (C) multiple times to obtain the compound shown below:
H2N—CH(R)—CO—(HN—CH(R)—CO)n—SH  (4)
(wherein each R, each independently, represents the side chain of any amino acid, and n represents in integer that is 2 or more).
[5] The manufacturing method according to [3] or [4], wherein said one or more third amino acid derivative(s) does not possess a protecting group.
[6] The manufacturing method according to any of [1] to [5], further comprising:

(D) a step of reacting Compound (2) obtained in step (B), or Compound (3) or Compound (4) obtained in step (C), with a nucleophilicity reagent shown below:
H—X  (5)
(wherein X represents a substituent bound via an atom selected from the group consisting of N, O, S, and Se)
to obtain the compound shown below:
H2N—CH(R)—CO—(HN—CH(R)—CO)m—X  (6)
(wherein each R, each independently, represents the side chain of any amino acid, m represents any integer, and X represents a substituent bound via an atom selected from the group consisting of N, O, S, and Se).
[7] The manufacturing method according to [6], wherein X is a substituent bound via an N atom or S atom.
[8] The manufacturing method according to [4], which is a method for manufacturing a polymer comprising 4 to 20 amino acids.
[9] The manufacturing method according to any of [1] to [8], wherein the first or second amino acid derivative, or at least one of said more than one third amino acid derivatives has an unprotected 4-aminobutyl group as a side chain.
[10] The manufacturing method according to any of [1] to [8], wherein the side chains of said first and second amino acid derivatives, and of said one or more third amino acid derivative are identical.
[11] The manufacturing method according to [10], wherein said side chains are all methyl groups.
[12] A method for manufacturing a polymer of more than one amino acids linked by peptide bonds, characterized in that it comprises:

(a) a step of preparing more than one amino acid derivatives shown below:
H2N—CH(R)—COSH  (1)
(wherein R represents the side chain of any amino acid);

(b) a step of subjecting said more than one amino acid derivatives to an oxidation reaction to obtain more than one compound shown below:
H2N—CH(R)—CO—(HN—CH(R)—CO)x—X—SH  (7)
(wherein each R, each independently, represents the side chain of any amino acid, x represents any integer)
and/or the compound shown below:
H2N—CH(R)—CO—(HN—CH(R)—CO)y—S—S—(CO—CH(R)—NH)z—CO—CH(R)—NH2(8)
(wherein each R, each independently, represents the side chain of any amino acid, and y and z represent any integer),

where a portion or all of said more than one amino acid derivatives do not possess a protecting group.

[13] The method according to [12], wherein all of said more than one amino acid derivatives do not possess a protecting group.

[14] The method according to [12] or [13], wherein the compound obtained in step (b) comprises Compound (7).

[15] The manufacturing method according to [14], further comprising:

(c1) a step of reacting Compound (7) obtained in step (b) with a nucleophilicity reagent shown below:
H—X  (5)
(wherein X represents a substituent bound by an atom selected from the group consisting of N, O, S, and Se)
to obtain the compound shown below:
H2N—CH(R)—CO—(HN—CH(R)—CO)x—X  (9)
(wherein each R, each independently, represents the side chain of any amino acid, x represents any integer, and X represents a substituent bound by an atom selected from the group consisting of N, O, S, and Se).
[16] The manufacturing method according to [15], wherein X is a substituent bound by an N atom or S atom.
[17] The manufacturing method according to [12] or [13], wherein the compound obtained in step (b) comprises Compound (8).
[18] The manufacturing method according to [17], further comprising:

(c2) a step of reacting Compound (8) obtained in step (b) with a nucleophilicity reagent shown below:
H—X  (5)
(wherein X represents a substituent bound by an atom selected from the group consisting of N, O, S, and Se)
to obtain the compound shown below:
H2N—CH(R)—CO—(HN—CH(R)—CO)y—X  (10)
(wherein each R, each independently, represents the side chain of any amino acid, y represents any integer, and X represents a substituent bound by an atom selected from the group consisting of N, O, S, and Se)
and below:
H2N—CH(R)—CO—(HN—CH(R)—CO)z—X  (11)
(wherein each R, each independently, represents the side chain of any amino acid, z represents any integer, and X represents a substituent bound by an atom selected from the group consisting of N, O, S, and Se).
[19] The manufacturing method according to [18], wherein X is a substituent bound by an N atom or S atom.
[20] The manufacturing method according to any of [12] to [19], which is a method for manufacturing a polymer comprising 4 to 20 amino acids.
[21] The manufacturing method according to any of [12] to [20], wherein at least one of said more than one amino acid derivatives has an unprotected 4-aminobutyl group as a side chain.
[22] The manufacturing method according to any of [12] to [21], wherein the side chains of said more than one amino acid derivatives are identical.
[23] The manufacturing method according to [22], wherein said side chains are all methyl groups.
[24] The manufacturing method according to any of [12] to [23] which is for producing a polypeptide library.

Those skilled in the art shall recognize that an invention of any combination of one or more characteristics of the present invention described above is encompassed by the scope of the present invention.

Effects of the Invention

According to the manufacturing method of the present invention, amino acid polymers can be manufactured extremely efficiently compared to conventional methods of utilizing a protecting group.

Moreover, since there is no need to utilize a protecting group in the manufacturing method of the present invention, problems involved with conventional methods that use highly toxic toxins for deprotection can be easily avoided.

Since the manufacturing method of the present invention allows synthesis of large amounts of more than one amino acid polymers, it may be suitable for industrial production of polyamino acids or production of a polypeptide library.

