Choline-binding domain as a novel affinity tag for purification of fusion proteins produced in Pichia pastoris

被引：15

作者：

Caubín, J

Martín, H

Roa, A

Cosano, I

Pozuelo, M

de la Fuente, JM

Sánchez-Puelles, JM

Molina, M

Nombela, C

机构：

[1] Univ Complutense, Fac Farm, Dept Microbiol 2, E-28040 Madrid, Spain

[2] SmithKline Beecham SA, Ctr Invest Basica, Tres Cantos, Spain

来源：

BIOTECHNOLOGY AND BIOENGINEERING | 2001年 / 74卷 / 02期

关键词：

C-LYTA; affinity tag; heterologous expression; yeast;

D O I：

10.1002/bit.1106

中图分类号：

Q81 [生物工程学（生物技术）]; Q93 [微生物学];

学科分类号：

071005 ; 0836 ; 090102 ; 100705 ;

摘要：

The choline-binding domain (ChoBD) of the carboxy-terminal region of the Streptococcus pneumoniae amidase LYTA (C-LYTA) presents a strong affinity for tertiary amines. We report a method for single-step purification of proteins expressed in the methylotrophic yeast Pichia pastoris based on the fusion of C-LYTA to the protein of interest. We show that C-LYTA can be efficiently expressed and secreted in this host. Tagged proteins fused to this binding domain can be purified on inexpensive DEAE matrices. It therefore provides a useful system for the purification of recombinant proteins with high specificity suitable for industrial purposes. (C) 2001 John Wiley & Sons, Inc.

引用

收藏

页码：164 / 171

页数：8

相关论文

共 20 条

[11] YEAST SHUTTLE AND INTEGRATIVE VECTORS WITH MULTIPLE CLONING SITES SUITABLE FOR CONSTRUCTION OF LACZ FUSIONS [J].

MYERS, AM ;

TZAGOLOFF, A ;

KINNEY, DM ;

LUSTY, CJ .

GENE, 1986, 45 (03) :299-310

[12] NOVEL METHOD FOR DETECTION OF BETA-LACTAMASES BY USING A CHROMOGENIC CEPHALOSPORIN SUBSTRATE [J].

OCALLAGHAN, CH ;

SHINGLER, AH ;

KIRBY, SM ;

MORRIS, A .

ANTIMICROBIAL AGENTS AND CHEMOTHERAPY, 1972, 1 (04) :283-+

[13] SINGLE-STEP PURIFICATION ON DEAE-SEPHACEL OF RECOMBINANT POLYPEPTIDES PRODUCED IN ESCHERICHIA-COLI [J].

ORTEGA, S ;

GARCIA, JL ;

ZAZO, M ;

VARELA, J ;

MUNOZWILLERY, I ;

CUEVAS, P ;

GIMENEZGALLEGO, G .

BIO-TECHNOLOGY, 1992, 10 (07) :795-798

[14] Contribution of novel choline-binding proteins to adherence, colonization and immunogenicity of Streptococcus pneumoniae [J].

Rosenow, C ;

Ryan, P ;

Weiser, JN ;

Johnson, S ;

Fontan, P ;

Ortqvist, A ;

Masure, HR .

MOLECULAR MICROBIOLOGY, 1997, 25 (05) :819-829

[15]

Sambrook J., 2002, MOL CLONING LAB MANU

[16] CLONING AND EXPRESSION OF GENE FRAGMENTS ENCODING THE CHOLINE-BINDING DOMAIN OF PNEUMOCOCCAL MUREIN HYDROLASES [J].

SANCHEZPUELLES, JM ;

SANZ, JM ;

GARCIA, JL ;

GARCIA, E .

GENE, 1990, 89 (01) :69-75

[17] IMMOBILIZATION AND SINGLE-STEP PURIFICATION OF FUSION PROTEINS USING DEAE-CELLULOSE [J].

SANCHEZPUELLES, JM ;

SANZ, JM ;

GARCIA, JL ;

GARCIA, E .

EUROPEAN JOURNAL OF BIOCHEMISTRY, 1992, 203 (1-2) :153-159

[18] STRUCTURAL REQUIREMENTS OF CHOLINE DERIVATIVES FOR CONVERSION OF PNEUMOCOCCAL AMIDASE - A NEW SINGLE-STEP PROCEDURE FOR PURIFICATION OF THIS AUTOLYSIN [J].

SANZ, JM ;

LOPEZ, R ;

GARCIA, JL .

FEBS LETTERS, 1988, 232 (02) :308-312

[19] HIGH-FREQUENCY TRANSFORMATION OF YEAST - AUTONOMOUS REPLICATION OF HYBRID DNA-MOLECULES [J].

STRUHL, K ;

STINCHCOMB, DT ;

SCHERER, S ;

DAVIS, RW .

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 1979, 76 (03) :1035-1039

[20] THE PUC PLASMIDS, AN M13MP7-DERIVED SYSTEM FOR INSERTION MUTAGENESIS AND SEQUENCING WITH SYNTHETIC UNIVERSAL PRIMERS [J].

VIEIRA, J ;

MESSING, J .

GENE, 1982, 19 (03) :259-268