Ajinomoto Co., Ltd.

Masayo Date

Keiichi Yokoyama

Yoshimi Kikuchi

Yukiko Umezawa

Daisuke Ejima

Hiroshi Matsui

Norimasa Onishi

Yoshiyuki Takahara

Hiroshi Itaya

Mototaka Suzuki

Haruna Sato

Akira Mitsui

Manabu Kitazawa

Yutaka Sato

Tatsuki Kashiwagi

Eiko Oshimura

Takami Maekawa

Eiichiro Suzuki

Sho Senda

Nami Nakamura

Yoko Kuriyama

Haruo Heima

Tomomi Yoshida

Seiichi Taguchi

Hisao Fukuda

Kouhei Tsumoto

Keisuke Kato

Yuichi Nakahara

Mayumi Watanabe

Yumi Ando

Takayoshi Fujii

The present invention provides a method for producing a refolded protein, the method comprising a step for mixing, inside a micro mixer, a buffer and a solubilization solution for a protein that has lost higher order structure or that has become insoluble.

Method for producing refolded protein by using flow microreactor, and protein refolding apparatus

Culture media, which contain albumin carrying a reduced amount of fatty acid, are useful for culturing stem cells.

Medium for culturing stem cells

A method for perming or coloring hair, in which hair is treated with a cosmetic composition including a protein-glutaminase and not including an alkaline substance. A method of enhancing hair-permeation of an active ingredient included in a cosmetic composition, in which a protein glutaminase is included in the cosmetic composition. The cosmetic composition does not include an alkaline substance, and the active ingredient is at least one of a hair perm agent and a hair coloring agent.

Cosmetic composition

The present invention provides a method for producing a multimeric protein composed of a monomeric protein, wherein the monomeric protein is obtained by fusing a protein having an immunoglobulin fold structure to a protein that can serve as a subunit structure, the method including the steps of:

Method for preparing polymeric protein composed of monomeric protein produced by fusing protein having immunoglobulin fold structure to protein capable of serving as subunit structure

A transglutaminase protein which is mutated to have improved heat resistance and/or pH stability. A mutation is introduced into WT transglutaminase at a cysteine residue capable of forming a disulfide bond (SS bond).

Transglutaminase having disulfide bond introduced therein

The present invention provides a method for efficiently producing an industrially useful protein in coryneform bacteria, and more particularly, a method for efficiently producing a protein for which secretion was difficult using conventional protein secretion pathways. In particular, the present invention provides a method for efficiently producing heterologous proteins comprising culturing coryneform bacteria containing an genetic construction containing a promoter sequence which functions in coryneform bacteria, a nucleic acid sequence encoding a Tat system-dependent signal peptide region, and a nucleic acid sequence encoding a heterologous protein, in the direction from 5'-end to 3&#x2032;-end, and secretory producing the heterologous protein by coryneform bacteria.

Method of producing proteins

The present invention provides transglutaminases with improved heat resistance. Specifically, the present invention provides mutant transglutaminase proteins with improved heat resistance as obtained by introducing appropriate mutations into transglutaminases, which results in the incorporation of a disulfide bond.

Thermotolerant transglutaminase originating in actinomyces

By culturing a methanol-assimilating bacterium which harbors a DNA construct which contains a promoter sequence that functions in the methanol-assimilating bacterium and a nucleotide sequence that encodes a polypeptide containing a signal sequence and an objective protein which is functionally connected to the promoter sequence, in a liquid medium containing methanol as a major carbon source, the bacterium is allowed to secrete the objective protein, and the secreted objective protein is recovered.

Method for producing protein

The present invention provides a neutral metalloprotease from actinomycetes which selectively cleaves a pro-structure part of a microbial protransglutaminase and a gene encoding said neutral metalloprotease. An active microbial transglutaminase having the pro-structure part cleaved can be obtained by culturing a microorganism into which a gene encoding the neutral metalloprotease from actinomycetes according to the present invention has been introduced, where by producing the neutral metalloprotease from actinomycetes, and reacting it on a microbial protransglutaminase.

Method of producing microbial transglutaminase

The present invention relates to a process for secretory production of a foreign protein, in particular, transglutaminase by a coryneform bacterium. According to the present invention, a process is provided for the secretory production of a foreign protein, in particular, transglutaminase, by making a coryneform bacterium to produce an industrially useful foreign protein, in particular, transglutaminase and efficiently release the product extracellularly (i.e., secretory production). An intended foreign protein, in particular, transglutaminase, is produced by using an expression construct wherein the gene sequence of the intended foreign protein containing the pro-structure part, in particular, pro-transglutaminase gene sequence, is ligated to the downstream of a sequence encoding the signal peptide region from a coryneform bacterium, introducing this expressional genetic construct into a coryneform bacterium, culturing the thus transformed coryneform bacterium, and treating the extracellularly released protein with a protease, etc. to cleave and eliminate the pro-part.

Process for producing transglutaminase

The object of the present invention is to provide a method of producing a heterologous protein by making a coryneform bacterium to produce and efficiently extracellularly secrete (secreto-production) an industrially useful heterologous protein. According to the present invention, a genetic construct is used where a gene sequence encoding an intended protein which is ligated to the downstream of a sequence encoding the signal peptide derived from a coryneform bacterium, the gene construct is introduced into a mutant coryneform bacterium which has a capacity of secreting the heterologous protein at least 2-fold higher than the wild typeATCC 13869, the mutant coryneform bacterium is cultured and the extracellularly released heterologous protein is recovered.

Methods for secretory production of proteins

In gene expression frequency analysis, a vector primer to which each cDNA is ligated is formed by synthesizing the cDNA from each mRNA using by a vector primer having a poly(T) sequence; each cDNA sequence is converted to a tag on the vector primer; a concatemer is formed by ligating the obtained tags via a sequence that enables recognition of ends of the tags; and the nucleotide sequence of the concatemer is analyzed.

Method for analyzing gene expression frequency

There are disclosed (i) a purification process for obtaining a human BCDF having the intramolecular disulfide linkage and the stereostructure of natural type human BCDF which comprises subjecting to an oxidation reaction and a refolding treatment a reduced type human BCDF obtained by culturing a microorganism having a human BCDF gene integrated therein and solubilized with guanidine hydrochloride, characterized in that after the oxidation reaction, a gel filtration chromatographic treatment is conducted under the conditions of the guanidine hydrochloride concentration adjusted to 4-7M; (ii) a purification process for obtaining a natural type human BCDF monomer by removing the organic solvent from an organic solvent-containing solution of human BCDF, characterized in that the solution is passed through a gel filtration chromatographic column equilibrated with an organic solvent, followed by eluting according to a stepwise or linear gradient program; and (iii) a human BCDF purification process comprising an ion exchange chromatographic treatment and a reversed phase HPLC step, in combination (i) or (ii). According to these purification processes, it becomes possible to remove the impurities derived from a microorganism and human BCDF analogs, and the thus obtained natural type human BCDF has a high purity and can be utilized for pharmaceutical preparations.

Growing community of inventors

Kawasaki, Japan

Masayo Date