公开(公告)号：US06635458B2

公开(公告)日：2003-10-21

申请号：US09308683

申请日：1999-07-15

申请人： Jose Luis Garcia Lopez ,  Estrella Cortes Rubio ,  Jorge Alonso Palacios ,  Encarnacion Mellado Duran ,  Bruno Diez Garcia ,  Jose Manuel Guisan Seijas ,  Franciso Salto Maldonado ,  Barredo Fuente

发明人： Jose Luis Garcia Lopez ,  Estrella Cortes Rubio ,  Jorge Alonso Palacios ,  Encarnacion Mellado Duran ,  Bruno Diez Garcia ,  Jose Manuel Guisan Seijas ,  Franciso Salto Maldonado ,  Barredo Fuente

IPC分类号： C12N904

CPC分类号： C12N9/0024 ,  C12P35/06

摘要： Enzymatic process for the preparation of cephalosporanic 7&bgr;-(4-carboxybutanamide) acid by using the modified enzyme D-aminoacid oxidase of Trigonopsis variabilis produced in Escherichia coli. The process for the expression of the enzyme comprises: (I) isolating the DNA corresponding to gene which codes for the enzyme D-aminoacid oxidase; (II) removing the intron which is contained in said gene; (III) inserting the DNA fragment obtained into the plasmide which is capable of replication in Escherichia coli; (IV) fusing at the extremity 5′ of the structural region of the gene a synthetic assembler which contains a nucleotide sequence which codes for six histidines; (V) transforming a strain of Escherichia coli with the resulting recombinant plasmide; (VI) cultivating the transformed cells of Escherichia coli; and (VII) recovering the enzyme D-aminoacid oxidase of the former cultivation operation through affinity chromatography.

摘要翻译： 通过使用在大肠杆菌中产生的变异型变异酶的修饰酶D-氨基酸氧化酶来制备头孢菌素7beta-（4-羧基丁酰胺）酸的酶法。 酶的表达过程包括：（I）分离与编码D-氨基酸氧化酶酶的基因相对应的DNA; （II）除去所述基因中所含的内含子; （III）将获得的DNA片段插入能在大肠杆菌中复制的质粒; （IV）在基因结构区域的末端5'处融合合成汇编器，其包含编码六个组氨酸的核苷酸序列; （V）用所得重组质粒转化大肠杆菌菌株; （VI）培养大肠杆菌的转化细胞; 和（VII）通过亲和层析回收前一种培养操作的酶D-氨基酸氧化酶。

公开(公告)号：US6146871A

公开(公告)日：2000-11-14

申请号：US981321

申请日：1998-08-13

申请人： Jose Luis Garcia Lopez ,  Estrella Cortes Rubio ,  Jose Manuel Guisan Seijas ,  Jose Luis Barredo Fuente ,  Bruno Diez Garcia ,  Alfonso Collados de la Vieja ,  Alejandro Vitaller Alba ,  Francisco Salto Maldonado

发明人： Jose Luis Garcia Lopez ,  Estrella Cortes Rubio ,  Jose Manuel Guisan Seijas ,  Jose Luis Barredo Fuente ,  Bruno Diez Garcia ,  Alfonso Collados de la Vieja ,  Alejandro Vitaller Alba ,  Francisco Salto Maldonado

IPC分类号： C12N15/09 ,  C12N1/21 ,  C12N9/78 ,  C12N9/84 ,  C12N9/86 ,  C12N15/00 ,  C12N15/55 ,  C12P17/14 ,  C12R1/01 ,  C12R1/19 ,  C12N9/18 ,  C12N15/70

CPC分类号： C12N9/86 ,  C12N9/84 ,  C07K2319/00

摘要： A procedure for producing a modified 7.beta.-(4-carboxybutanamide) cephalosporinase enzyme which can be purified in a single chromatographic step. The procedure for production of the enzyme involves: mutagenizing the gene which codes for the enzyme from Acinetobacter sp. ATCC 53891 by inserting a nucleotide sequence coding for six histidine residues; fusing the mutant gene with high-efficiency promoter DNA sequences; transforming Escherichia coli cells with the fusion gene construct; growing the transformed Escherichia coli cells; and recovering the enzyme by the use of supports which contain metal chelates. 7-Aminocephalosporanic acid is an important intermediate for the manufacture of a wide range of antibacterial agents of the cephalosporin family.

摘要翻译： PCT No.PCT / ES97 / 00098 Sec。 371日期1998年8月13日 102（e）日期1998年8月13日PCT 1997年4月18日PCT PCT。 公开号WO97 / 40175 PCT 日期1997年10月30日制备修饰的7β-（4-羧基丁酰胺）头孢菌素酶的方法，其可以在单个色谱步骤中纯化。 酶的制备方法包括：从不动杆菌属（Acinetobacter sp。）中诱变编码酶的基因。 ATCC 53891通过插入编码六个组氨酸残基的核苷酸序列; 将突变基因与高效启动子DNA序列融合; 用融合基因构建体转化大肠杆菌细胞; 生长转化的大肠杆菌细胞; 并通过使用含有金属螯合物的载体回收酶。 7-氨基头孢烷酸是制造头孢菌素类的广泛抗菌剂的重要中间体。

公开(公告)号：US06187574B1

公开(公告)日：2001-02-13

申请号：US08973914

申请日：1998-04-28

申请人： Jose Luis Garcia Lopez ,  Estrella Cortes Rubio ,  Jorge Alonso Palacios ,  Jose Luis Barredo Fuente ,  Bruno Diez Garcia ,  Miguel Angel Moreno Valle ,  Carmen Schleissner Sanchez ,  Alfonso Collados de la Vieja ,  Alejandro Vitaller Alba ,  Francisco Salto Maldonado

发明人： Jose Luis Garcia Lopez ,  Estrella Cortes Rubio ,  Jorge Alonso Palacios ,  Jose Luis Barredo Fuente ,  Bruno Diez Garcia ,  Miguel Angel Moreno Valle ,  Carmen Schleissner Sanchez ,  Alfonso Collados de la Vieja ,  Alejandro Vitaller Alba ,  Francisco Salto Maldonado

IPC分类号： C12N902

CPC分类号： C12N9/0024

摘要： Process for producing the enzyme D-amino acid oxydase of Rhodotorula gracilis in host cells. The process for the expression of the enzyme comprises isolating the complementary DNA corresponding to the messager RNA of the gene which codes for the D-amino acid oxydase of any strain of Rhodotorula gracillis producing said enzyme; fusing the fragment of DNA which codes for D-amino acid oxydase of Rhodotorula gracillis with a DNA sequence which contains a site of union to the ribosome and a high efficiency promoter sequence for the express of genes in host cells; inserting the DNA fragment into a plasmid appropriate for the host cell; cultivating the host cells transformed with said plasmid; and collecting the enzyme.

摘要翻译： 在宿主细胞中生产细小红球菌的D-氨基酸氧化酶酶的方法。 表达酶的过程包括分离与编码产生所述酶的任何红曲霉菌株的D-氨基酸氧化酶的基因的信使RNA相对应的互补DNA; 将编码红景天细菌的D-氨基酸氧化酶的DNA片段与包含与核糖体结合的位点的DNA序列和用于在宿主细胞中表达基因的高效启动子序列融合; 将DNA片段插入适合宿主细胞的质粒中; 培养用所述质粒转化的宿主细胞; 并收集酶。

公开(公告)号：US20130274510A1

公开(公告)日：2013-10-17

申请号：US13825447

申请日：2011-09-20

申请人： Isabel Fernandez Escapa ,  Ma del Valle Morales Ruiz ,  Jose Luis Garcia Lopez ,  Ma Auxiliadora Prieto Jimenez

发明人： Isabel Fernandez Escapa ,  Ma del Valle Morales Ruiz ,  Jose Luis Garcia Lopez ,  Ma Auxiliadora Prieto Jimenez

IPC分类号： C12P7/62 ,  C07C323/52 ,  C12N15/78 ,  C07C319/20 ,  C08G63/78 ,  C12P11/00

CPC分类号： C12P7/625 ,  C07C319/20 ,  C07C323/52 ,  C07C327/32 ,  C08G63/78 ,  C12N9/0006 ,  C12N15/78 ,  C12P11/00 ,  C12R1/40

摘要： The present invention relates to a process for the synthesis of polyhydroxyalkanoates (PHAs) and PHAs obtained by the said process and the use of the said PHAs. The PHAs of the present invention, called PHACOS, have a novel monomeric composition, as they contain monomers with thioester groups in the side chain that confer new physicochemical properties and allow subsequent chemical modification.

摘要翻译： 本发明涉及通过所述方法获得的聚羟基链烷酸酯（PHA）和PHA的合成方法和所述PHA的使用。 称为PHACOS的本发明的PHA具有新颖的单体组成，因为它们含有在侧链中具有硫酯基团的单体，赋予新的物理化学性质并允许随后的化学改性。