The present invention provides, as a novel compound that can be utilized for efficient synthesis or the like of a polypeptide comprising various amino acids, a silane-containing condensed cyclic dipeptide compound represented by formula (A).

##STR00001##

In formula (A), each of R.sup.11, R.sup.12, R.sup.13, R.sup.21, and R.sup.22 independently represents a hydrogen atom, a halogen atom, a hydroxyl group, a carboxyl group, a nitro group, a cyano group, or a thiol group, or a monovalent aliphatic hydrocarbon group, aromatic hydrocarbon group, or heterocyclic group that may have one or more substituents; and each of R.sup.a1 and R.sup.a2 independently represents a monovalent aliphatic hydrocarbon group or aromatic hydrocarbon group that may have one or more substituents

Processes for performing peptide synthesis, particularly for cyclic peptide synthesis are generally described. Reaction intermediates of the peptide synthesis are also described.

The invention relates to a method for enzymatically synthesizing an (oligo)peptide, comprising coupling (a) an (oligo)peptide C-terminal ester or thioester and (b) an (oligo)peptide nucleophile having an N-terminally unprotected amine, wherein the coupling is carried out in a fluid comprising water, and wherein the coupling is catalyzed by a subtilisin BPN variant or a homolog thereof, which comprises the following mutations compared to subtilisin BPN represented by SEQUENCE ID NO: 2 or a homolog sequence thereof: a deletion of the amino acids corresponding to positions 75-83; a mutation at the amino acid position corresponding to S221, the mutation being S221C or S221 selenocysteine; preferably a mutation at the amino acid position corresponding to P225 wherein the amino acid positions are defined according to the sequence of subtilisin BPN represented by SEQUENCE ID NO: 2. Further, the invention relates to an enzyme suitable for use as a catalyst in a method of the invention.

The invention relates to a subtilisin BPN variant or homologue thereof, having the following mutations compared to subtilisin BPN represented by SEQUENCE ID NO: 2 or a homologue sequence thereof:a deletion of the amino acids corresponding to positions 75-83;a mutation at the amino acid position corresponding to S221, the mutation corresponding to S221C or S221 selenocysteine, preferably S221C;at least one further mutation selected from the group consisting of amino acid positions corresponding to F189W, F189Y, S33D, S33T, N218D, N218T, N218E, N62D, N62S, N62W, and N62Y; preferably a mutation at the amino acid position corresponding to P225; wherein the amino acid positions are defined according to the sequence of subtilisin BPN represented by SEQUENCE ID NO: 2. The invention further relates to enzymatically synthesizing a peptide.

The invention relates to a method for enzymatically synthesizing an (oligo)peptide, comprising coupling (a) an (oligo)peptide C-terminal ester or thioester and (b) an (oligo)peptide nucleophile having an N-terminally unprotected amine,

wherein the coupling is carried out in a fluid comprising water, and

wherein the coupling is catalyzed by a subtilisin BPN variant or a homologue thereof, which comprises the following mutations compared to subtilisin BPN represented by SEQUENCE ID NO: 2 or a homologue sequence thereof :

a deletion of the amino acids corresponding to positions 75-83;

a mutation at the amino acid position corresponding to S221, the mutation being S221C or S221 selenocysteine;

preferably a mutation at the amino acid position corresponding to P225 wherein the amino acid positions are defined according to the sequence of subtilisin BPN represented by SEQUENCE ID NO: 2.

Further, the invention relates to an enzyme suitable for use as a catalyst in a method of the invention.

Solid supports for use in solid-phase peptide synthesis (SPPS) are provided. The solid supports may include a resin and a protected linker coupled to the resin. The linker may be an N-mercaptoethoxyglycine, an N-mercaptopropoxyglycine, an N-mercaptobutoxyglycine, and/or another suitable linker. Kits for use in SPPS are also provided. The kits may include a solid support, a solution including a thiol or a selenol, one or more pluralities of protected amino acids, and/or a wash buffer. Methods of SPPS are also provided. The methods may include providing a solid support including a resin coupled to a protected linker.

A method of downstream purification of a recombinant protein in fusion with a hydrophobin-intein tag using alcohol precipitation. The method comprises (1) constructing a plasmid expressing a fusion protein in a host cell, wherein the fusion protein includes a target protein domain, an intein, and a hydrophobin domain; (2) culturing the host cell transfected with the plasmid forming a cell culture medium; (3) separating a cell culture supernatant containing the fusion protein from the cell culture medium; and (4) purifying a substantial amount of the target protein from the cell culture supernatant without its hydrophobin-intein fusion tag using alcohol precipitation.

The invention relates to a method for enzymatically synthesizing an (oligo)peptide, comprising coupling (a) an (oligo)peptide C-terminal ester or thioester and (b) an (oligo)peptide nucleophile having an N-terminally unprotected amine, wherein the coupling is carried out in a fluid comprising water, and wherein the coupling is catalyzed by a subtilisin BPN variant or a homo-log thereof, which comprises the following mutations compared to subtilisin BPN represented by SEQUENCE ID NO: 2 or a homolog sequence thereof: a deletion of the amino acids corresponding to positions 75-83; a mutation at the amino acid position corresponding to S221, the mutation being S221C or S221 selenocysteine; preferably a mutation at the amino acid position corresponding to P225 wherein the amino acid positions are defined according to the sequence of subtilisin BPN represented by SEQUENCE ID NO: 2. Further, the invention relates to an enzyme suitable for use as a catalyst in a method of the invention.

A preparation method for an N-long-chain acyl amino acid dipeptide, supramolecular amino acid and corresponding salts thereof, capable of controlling the occurrence of structural reconstruction and controlling the content of long-chain fatty acid. Further provided are a supramolecular amino acid and salts thereof, as well as an application in the fields of daily chemicals and the like.

The present disclosure relates to a novel spherical agglomeration method for proteins, and protein particles made by the spherical agglomeration method. By using continuous oscillatory baffled crystallizer, the method of the present disclosure is capable of maintain the biologically activities and providing protein particles with an average particle size between 1-500 ?m.