SPECIFIC INTERACTIONS OF ERWINIA-CHRYSANTHEMI KDGR REPRESSOR WITH DIFFERENT OPERATORS OF GENES INVOLVED IN PECTINOLYSIS

被引:99
作者
NASSER, W [1 ]
REVERCHON, S [1 ]
CONDEMINE, G [1 ]
ROBERTBAUDOUY, J [1 ]
机构
[1] INST NATL SCI APPL,GENET MOLEC MICROORGANISMES & INTERACT CELLULAIRE,CNRS,URA 1486,F-69621 VILLEURBANNE,FRANCE
关键词
PECTINOLYTIC BACTERIA; SOFT-ROT; PHYTOPATHOGEN; REGULATION; REPRESSOR-BINDING SITE;
D O I
10.1006/jmbi.1994.1155
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The Erwinia chrysanthemi kdgR gene encodes a repressor that negatively regulates the expression of genes involved in pectinolysis and in pectinase secretion. The cloned kdgR gene was overexpressed in Escherichia coli by using a phage T7 system. Overproduced repressor was purified to homogeneity by two chromatographic steps. Gel retardation and DNase I protection experiments demonstrated the specific binding of the KdgR protein to the operators of pectinase genes (pelA, pelB, pelC, pelE), to the operator of genes involved in pectin catabolism (kdgT, ogl, kduI-kdgF) and to that of the outT gene involved in pectinase secretion. These interactions involved one (pelA, pelB, kduI-kdgF, outT) or several operator sites (pelC, pelE, ogl, kdgT) that generally overlap the promoter. Despite the presence of potential KdgR binding sites (KdgR-box) in the regulatory regions of four genes involved in pectin catabolism (kdgC, kduD, pem, kdgA) and in a pectinase secretion gene outC, no DNA-repressor complex could be observed by in vitro experiments. By using a missing contact experiment on the coding strand of ogl and pelE regulatory regions, a new KdgR-binding consensus was proposed. This new consensus, constituted by two half motifs (AAT(A)(G)AAA(T)(C))N(N(CA)(TG)TTT(T)(C)A), is well conserved in the operators which interact in vitro with the KdgR repressor. In contrast, this repressor-recognized motif is degenerated in the other operators that cannot interact in vitro with the repressor. These results suggest the existence of different regulation mechanisms mediated by the KdgR protein for the two classes of operators. © 1994 Academic Press Limited.
引用
收藏
页码:427 / 440
页数:14
相关论文
共 42 条
[1]   NUCLEOTIDE-SEQUENCE OF THE ERWINIA-CHRYSANTHEMI GENE ENCODING 2-KETO-3-DEOXYGLUCONATE PERMEASE [J].
ALLEN, C ;
REVERCHON, S ;
ROBERTBAUDOUY, J .
GENE, 1989, 83 (02) :233-241
[2]   ATG VECTORS FOR REGULATED HIGH-LEVEL EXPRESSION OF CLONED GENES IN ESCHERICHIA-COLI [J].
AMANN, E ;
BROSIUS, J .
GENE, 1985, 40 (2-3) :183-190
[3]  
Ausubel FM, 1987, CURRENT PROTOCOLS MO
[4]   LINKAGE MAP OF ESCHERICHIA-COLI-K-12, EDITION-7 [J].
BACHMANN, BJ .
MICROBIOLOGICAL REVIEWS, 1983, 47 (02) :180-230
[6]   REGULATION AND ROLE IN PATHOGENICITY OF ERWINIA-CHRYSANTHEMI 3937 PECTIN METHYLESTERASE [J].
BOCCARA, M ;
CHATAIN, V .
JOURNAL OF BACTERIOLOGY, 1989, 171 (07) :4085-4087
[7]  
BOURSON C, 1993, J GEN MICROBIOL, V139, P1
[8]  
BOURSON C, 1992, THESIS C BERNARD U L
[9]  
BRADFORD MM, 1976, ANAL BIOCHEM, V72, P248, DOI 10.1016/0003-2697(76)90527-3
[10]   MISSING CONTACT PROBING OF DNA-PROTEIN INTERACTIONS [J].
BRUNELLE, A ;
SCHLEIF, RF .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 1987, 84 (19) :6673-6676