• 专利附录
  • 专利说明
  • 权利要求
  • 类似技术
  • 同族专利
  • 引用文献
  • 法律状态
  • 优先权
    • 专利摘要:
    Human insulin is semi-synthetically prepared using a novel method comprising the preparation of a modified human insulin of the general formula …<CHEM>… wherein R = H or an enzymatically separable sugar residue or an ester thereof, and R<1> = an enzymatically separable sugar residue or an ester thereof, or an amino acid or peptide, or an amide or ester of the latter two, or R<1> - OH if R NOTEQUAL H, followed by enzymatic removal of the modifying groups R and/or R<1> to yield human insulin.
    公开号:SU1232148A3
    申请号:SU833582755
    申请日:1983-04-20
    公开日:1986-05-15
    发明作者:Виллем Карел Ван Дедем Гейсберт；Эгберт Абрахам Ван Хауденховен Франсуа
    申请人:Акцо Н.В. (Фирма)；
    IPC主号:
    专利说明:


    This invention relates to biochemistry, in particular, to obtaining insulin tracer by semisynthetic means.
    The purpose of the invention is to increase the flow rate and simplify the process for obtaining human insulin by using modified insulin of the general formula.
    Des (VZO) -insulin-NH-CH-C-R
    ns-ns
    I
    he


    where R is phenylalanyl-phenylalanine, which is subjected to the processing of carboxy- | 5
    by separatidase A, specifically cleaved-β-phe-phe was isolated by chromatography. Example 2. DAI was dissolved in medium consisting of 60% of the organic fraction (see Example 1) and 40% of 0.5 M Tris-HC1 buffer to a concentration of 32.2 g / l. Then, the thr-phe-phe-HCl tripeptide is dissolved in this medium to a concentration of 0.047 M. With. A 0.1 M solution of NaOH adjusted the pH of the mixture to 6.5. After that, immobilization on trypsin on silica gel is introduced into the solution up to a concentration of 157 g / l. The reaction mixture is incubated at 37 ° C for 3 hours. DAI throne R to form human insulin.
    The method is carried out as follows.
    Example 1. As a starting compound, des (VZO) - - stroke (DAI), obtained from pig insulin according to the Schmitt method, is used. First, DAI is bound to the thr-phe-phe tripeptide. A mixture consisting of 67% of the organic fraction (dimethylformamide and ethanol 1: 1) and 33% of 0.5 M Tris-HC1 buffer are used as the reaction medium. Dai is dissolved in this medium to a concentration of 13.6 g / l and tripeptide is weighed to a concentration of 0.067 M. The mixture is adjusted to a pH of 6.5 with 0.1 M NaOH solution. Trypsin is then added to the solution.
    to a concentration of 5 g / l and the reaction mixture is incubated for 35 min. DAI-thr-phe-phe was separated from the reaction system by chromatography and lyophilized. The yield is 54%. DAI-thr-phe-phe is dissolved in 0.2 M ammonium bicarbonate solution from pH 8.5 to 20
    thirty
    method and subjected to lyophilization. The yield of the drug is 27%; The conversion of DAI-thr-phe-phe into human insulin and the insulin investing are carried out as in Example 1.
    Example 3. DAI was dissolved in the medium prepared according to Example 2 to a concentration of 25.2 g / l. Then, the tripeptide thr-phe-phe-HCl is dissolved to a concentration of 0.037 M. The pH of the mixture is adjusted to 6.3 with 0.1 M NaOH. After that, the lysyl endopeptidase immobilized on silica gel is introduced into the solution to a concentration of 77 g / l, and the reaction mixture is incubated at 37 ° C for 48 h. The yield of the drug is 38%. The transformation of the tai-thr-phe-phe into human insulin and the administration of insulin is carried out according to example G.
    The identity of the obtained preparation with human insulin is confirmed by the data of amino acid analysis, indicating the complete correspondence of the amino acid composition of the preparation and insulin of humans
    40
    Ceitration 2.8 mg / ml. To this solution 45 ka. High purity (homogeneity)
    2 µl of (2,3 E) carboxypeptidase is added. The enzyme reaction is stopped after 30 minutes by adding an equal volume of 0.5 M citric acid solution. The resulting ins-phe-phe is isolated by chromatographic
    The human LNS is crystallized.
    Example 2. DAY is dissolved in medium consisting of 60% of the organic fraction (see Example 1) and 40% of 0.5 M Tris-HCl buffer to a concentration of 32.2 g / l. Then, the thr-phe-phe-HCl tripeptide is dissolved in this medium to a concentration of 0.047 M. With. A 0.1 M solution of NaOH adjusted the pH of the mixture to 6.5. After that, immobilization on trypsin on silica gel is introduced into the solution up to a concentration of 157 g / l. The reaction mixture is incubated at 37 ° C for 3 hours. DAI-thr-phe-phe is isolated by chromatographic
    0
    0
    method and subjected to lyophilization. The yield of the drug is 27%; The conversion of DAI-thr-phe-phe into human insulin and the insulin investing are carried out as in Example 1.
    Example 3. DAI was dissolved in the medium prepared according to Example 2 to a concentration of 25.2 g / l. Then, the tripeptide thr-phe-phe-HCl is dissolved to a concentration of 0.037 M. The pH of the mixture is adjusted to 6.3 with 0.1 M NaOH. After that, the lysyl endopeptidase immobilized on silica gel is introduced into the solution to a concentration of 77 g / l, and the reaction mixture is incubated at 37 ° C for 48 h. The yield of the drug is 38%. The transformation of the tai-thr-phe-phe into human insulin and the administration of insulin is carried out according to example G.
    The identity of the obtained preparation with human insulin is confirmed by the data of amino acid analysis, indicating the complete correspondence of the amino acid composition of the preparation and insulin of human 5
    0
    the human insulin preparation obtained was proven by high pressure liquid chromatography (one main peak with a retention time of 1089).
    权利要求:
    Claims (1)
    [1]
    METHOD FOR PRODUCING HUMAN INSULIN, characterized in that, in order to improve purity and simplify the method, modified human insulin of the general formula
    About des (HCO) -insulin-nh-CH-C-R ns-sn.
    I
    OH · where R is phenylalanyl-fenylalanine, subjected to treatment with carboxypeptidase A, followed by isolation of the target product.
    g with
    At · ”1232148 AZ 1 1232
    类似技术:
    公开号 | 公开日 | 专利标题
    Ståhl et al.1990|The synthesis of a D-amino acid ester in an organic media with α-chymotrypsin modified by a bio-imprinting procedure
    EP0020290B1|1984-07-11|Process for the specific splitting-off of protein sequences from proteins
    KR100885149B1|2009-02-23|Process for producing sugar chain asparagine derivative
    JP2994486B2|1999-12-27|Lipopeptide deacylase
    JP3194928B2|2001-08-06|Pre-expression method of gene in cell-free system of conjugation transcription / translation
    EP0003786B1|1980-12-10|Stereoselective separation of phenylglycine derivatives and 4-hydroxyphenylglycine derivatives with enzyme-containing polymers
    SU1232148A3|1986-05-15|Method of producing insulin of man
    Heuermann et al.1991|Purification and characterization of a sialidase fromClostridium chauvoei NC08596
    DE3626915A1|1988-02-11|METHOD FOR THE ENZYMATIC SYNTHESIS OF A TRISACCHARID CONTAINING N-ACETYLNEURAMINE ACID
    Neumann et al.1968|Intramolecular acyl migration in adenosine derivatives.
    DE2509482C2|1985-11-21|Process for the preparation of the kallikrein trypsin inhibitor from bovine lung
    EP0367302A2|1990-05-09|Process for semi-synthesis of human insulin, water-soluble cross-linked Achromobacter protease I for use therein and a process for preparing the same
    KATAGIRI et al.1959|ON THE TRANSGLYCOSIDATION RELATING TO RIBOFLAVIN BY ESCHERICHIA COLI III. ISOLATION OF TRANSGLUCOSIDASE AND FURTHER STUDIES ON THE SPECIFICITIES OF THE ENZYME
    US4532214A|1985-07-30|Method for isolation of aminoacylase
    JP2891023B2|1999-05-17|Method for producing lanthionine-containing material
    US4374197A|1983-02-15|Process for thymosin α1
    EP0059182B1|1984-11-21|A process for isolating aminoacylase enzyme from a mammal kidney extract
    US3716551A|1973-02-13|Deuterated l-amino acid mixture
    JPH05176785A|1993-07-20|Production of arbutin
    JPH0998779A|1997-04-15|Trehalose synthetase, its production and production of trehalose using the enzyme
    JP2690779B2|1997-12-17|L-ascorbic acid derivative and method for producing the same
    EP0518088A2|1992-12-16|Process for clostripain-catalysed coupling of Arg-Pro and Arg-B | containing them
    KATAGIRI et al.1959|ON THE TRANSGLYCOSIDATION RELATING TO RIBOFLAVIN BY ESCHERICHIA COLI
    JP2887689B2|1999-04-26|Method for producing glycyrrhetinic acid monoglucuronide
    SU455546A3|1974-12-30|Aspraginase release method
    同族专利:
    公开号 | 公开日
    AU1355483A|1983-10-27|
    IL68397D0|1983-07-31|
    IL68397A|1986-01-31|
    NL8201650A|1983-11-16|
    JPH0735399B2|1995-04-19|
    JPH05339290A|1993-12-21|
    PL143571B1|1988-02-29|
    AT42203T|1989-05-15|
    ZA832602B|1984-01-25|
    ES521674A0|1984-06-16|
    EP0092280B1|1989-04-19|
    US4840897A|1989-06-20|
    DK173007B1|1999-11-08|
    ES8405760A1|1984-06-16|
    AU554050B2|1986-08-07|
    JPS58189149A|1983-11-04|
    PL241575A1|1983-12-19|
    DK161683A|1983-10-22|
    HU189268B|1986-06-30|
    JPH0533993B2|1993-05-20|
    DK161683D0|1983-04-13|
    DE3379640D1|1989-05-24|
    CA1202587A|1986-04-01|
    EP0092280A1|1983-10-26|
    引用文献:
    公开号 | 申请日 | 公开日 | 申请人 | 专利标题
    US3276961A|1963-04-08|1966-10-04|Squibb & Sons Inc|Process for preparing human insulin|
    US4182654A|1974-09-18|1980-01-08|Pierce Chemical Company|Production of polypeptides using polynucleotides|
    JPS5718799B2|1979-04-13|1982-04-19|
    DK146482C|1979-04-13|1986-10-06|Shionogi & Co|METHOD FOR PREPARING A B30-THREONINE INSULIN|
    JPS5746360B2|1979-04-13|1982-10-02|
    US4343898A|1980-02-11|1982-08-10|Novo Industri A/S|Process for preparing esters of human insulin|
    DK147437A|1980-02-11|1900-01-01|Process for preparing human insulin or threonine B30 esters of human insulin, or a salt or complex thereof|
    DK319780A|1980-07-24|1982-01-25|Forenede Bryggerier As|PROCEDURE FOR ENZYMATIC REPLACEMENT OF B-30 AMINO ACID IN INSULINES|
    DE3327928A1|1983-08-03|1985-02-21|Hoechst Ag, 6230 Frankfurt|METHOD FOR PRODUCING INSULIN DERIVATIVES|DE3326472A1|1983-07-22|1985-02-14|Hoechst Ag, 6230 Frankfurt|NEW INSULIN DERIVATIVES, METHOD FOR THE PRODUCTION AND USE THEREOF AND PHARMACEUTICAL AGENTS FOR TREATING THE DIABETES MELLITUS|
    DE3326473A1|1983-07-22|1985-01-31|Hoechst Ag, 6230 Frankfurt|PHARMACEUTICAL AGENT FOR TREATING THE DIABETES MELLITUS|
    DE3327709A1|1983-07-29|1985-02-07|Hoechst Ag, 6230 Frankfurt|INSULIN DERIVATIVE CRYSTAL SUSPENSIONS, METHOD FOR THE PRODUCTION AND USE THEREOF|
    DE3333640A1|1983-09-17|1985-04-25|Hoechst Ag, 6230 Frankfurt|METHOD FOR THE PRODUCTION OF INSULIN DERIVATIVES, THE B-CHAIN C-TERMINAL EXTENDED, NEW BASICALLY MODIFIED INSULIN DERIVATIVES, THE MEANS CONTAINING THEM AND THEIR USE|
    DK309184D0|1984-06-25|1984-06-25|Nordisk Insulinlab|PROCEDURE FOR INSULATING INSULIN OR INSULINARY MATERIALS FROM A FERMENTING FLUID|
    法律状态:
    优先权:
    申请号 | 申请日 | 专利标题
    NL8201650A|NL8201650A|1982-04-21|1982-04-21|SEMISYNTHETIC PREPARATION OF HUMANE INSULIN.|
    [返回顶部]