Cloning, expression, purification, and characterization of recombinant flagellin isolated from Pseudomonas aeruginosa

被引：24
作者：
Goudarzi, Gholamreza [1 ]
Sattari, Morteza [1 ]
Roudkenar, Mehryar Habibi [2 ]
Montajabi-Niyat, Mehran [1 ]
Zavaran-Hosseini, Ahmad [3 ]
Mosavi-Hosseini, Kamran [2 ]
机构：
[1] Tarbiat Modares Univ, Dept Bacteriol, Sch Med Sci, Tehran, Iran
[2] Iranian Blood Transfus Org, Res Ctr, Tehran, Iran
[3] Tarbiat Modares Univ, Dept Immunol, Sch Med Sci, Tehran, Iran
来源：
BIOTECHNOLOGY LETTERS | 2009年 / 31卷 / 09期
关键词：
fliC; Inclusion bodies; Pseudomonas aeruginosa; Recombinant flagellin; PATHOGENESIS; GLYCOSYLATION;
D O I：
10.1007/s10529-009-0026-1
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：
Pseudomonas aeruginosa as an opportunistic pathogen causes lethal infections in immunocompromised individuals. This bacterium possesses a polar flagellum made up of flagellin subunits. Flagella have important roles in motility, chemotaxis, and establishment of P. aeruginosa in acute phase of infections. Isolation, cloning, and expression of flagellin were aimed at in this study. Flagellin gene (fliC) of P. aeruginosa strain 8821M was isolated by PCR and cloned into a pET expression vector. The recombinant flagellin (46 kDa) was overexpressed as inclusion bodies (IBs). IBs were solubilized in guanidine hydrochloride (GuHCl) followed by affinity-purification and renatured using Ni2+-Sepharose resin. Recombinant flagellins reacted with the serum from a rabbit previously immunized with native flagellin. In addition, polyclonal antiserum raised against the recombinant flagellin was shown to significantly inhibit the cell motility of P. aeruginosa strain 8821M in vitro.
引用
收藏
页码：1353 / 1360
页数：8
相关论文
共 23 条
[1]   ELECTROPHORETIC SEPARATION AND MOLECULAR-WEIGHT CHARACTERIZATION OF PSEUDOMONAS-AERUGINOSA H-ANTIGEN FLAGELLINS [J].
ALLISON, JS ;
DAWSON, M ;
DRAKE, D ;
MONTIE, TC .
INFECTION AND IMMUNITY, 1985, 49 (03) :770-774
[2]  
ANSORG R, 1978, ZENTBL BAKTERIOL PAR, V1, P228
[3]   Role of motility and flagellin glycosylation in the pathogenesis of Pseudomonas aeruginosa burn wound infections [J].
Arora, SK ;
Neely, AN ;
Blair, B ;
Lory, S ;
Ramphal, R .
INFECTION AND IMMUNITY, 2005, 73 (07) :4395-4398
[4]   Sequence polymorphism in the glycosylation island and flagellins of Pseudomonas aeruginosa [J].
Arora, SK ;
Wolfgang, MC ;
Lory, S ;
Ramphal, R .
JOURNAL OF BACTERIOLOGY, 2004, 186 (07) :2115-2122
[5]   ISOLATION AND CHARACTERIZATION OF PSEUDOMONAS-PSEUDOMALLEI FLAGELLIN PROTEINS [J].
BRETT, PJ ;
MAH, DCW ;
WOODS, DE .
INFECTION AND IMMUNITY, 1994, 62 (05) :1914-1919
[6]   Cloning and comparison of fliC genes and identification of glycosylation in the flagellin of Pseudomonas aeruginosa a-type strains [J].
Brimer, CD ;
Montie, TC .
JOURNAL OF BACTERIOLOGY, 1998, 180 (12) :3209-3217
[7]   Vaccines and immunotherapy against Pseudomonas aeruginosa [J].
Doering, Gerd ;
Pier, Gerald B. .
VACCINE, 2008, 26 (08) :1011-1024
[8]   A double-blind randomized placebo-controlled phase III study of a Pseudomonas aeruginosa flagella vaccine in cystic fibrosis patients [J].
Doering, Gerd ;
Meisner, Christoph ;
Stern, Martin .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2007, 104 (26) :11020-11025
[9]   The epidemiology, pathogenesis and treatment of Pseudomonas aeruginosa infections [J].
Driscoll, James A. ;
Brody, Steven L. ;
Kollef, Marin H. .
DRUGS, 2007, 67 (03) :351-368
[10]   Role of flagella in pathogenesis of Pseudomonas aeruginosa pulmonary infection [J].
Feldman, M ;
Bryan, R ;
Rajan, S ;
Scheffler, L ;
Brunnert, S ;
Tang, H ;
Prince, A .
INFECTION AND IMMUNITY, 1998, 66 (01) :43-51
123