The 5' region of cnf1 harbours a translational regulatory mechanism for CNF1 synthesis and encodes the cell-binding domain of the toxin.
The Escherichia coli cytotoxic necrotizing factor 1 (CNF1) is organized into three functional domains: the N-terminal part containing the cell-binding domain, a putative central membrane-spanning region, and a C-terminal catalytic region. On the basis of competition assays between CNF1 and GST-recombinant proteins containing different N-terminal fragments, and point mutations, we restricted the binding region to the first 190 amino acids. Hydrophilic amino acids 53-75 are strictly necessary to cell receptor recognition. Using different cnf1-lacZ translational fusions, we demonstrated that the mRNA corresponding to the first 48 codons of cnf1 is involved in the translational regulation of CNF1 synthesis. This regulation consists of both a positive and a negative control. The positive control is exerted by codons 6-20, including a putative downstream box that enhances the translational expression of cnf1. The negative control depends on codons 45-48. In this region, an anti-Shine-Dalgarno sequence, highly homologous to the core of the internal complementary sequence already reported for growth rate-regulated metabolic genes, has been detected. To some extent, the inner structural organization of CNF1 would thus suggest the compiling of several functions in a single mRNA protein system.
Fabbri
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INSERM Unité 452, Faculté de Médecine, 28 Avenue de Valombrose, F-06107, Nice, France.
Gauthier
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Boquet
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D020121:5' Untranslated Regions / Q000235:genetics*; D001427:Bacterial Toxins / Q000235:genetics*; D001665:Binding Sites; D002460:Cell Line / Q000378:metabolism; D003062:Codon / Q000235:genetics; D003603:Cytotoxins / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins; D015964:Gene Expression Regulation, Bacterial / Q000235:genetics*; D016297:Mutagenesis, Site-Directed; D014176:Protein Biosynthesis; D012329:RNA, Bacterial / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics; D012045:Regulatory Sequences, Nucleic Acid; D017384:Sequence Deletion; D012689:Sequence Homology, Nucleic Acid
D020121:5' Untranslated Regions; D001427:Bacterial Toxins; D003062:Codon; D003603:Cytotoxins; D029968:Escherichia coli Proteins; D012329:RNA, Bacterial; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; C060296:cytotoxic necrotizing factor type 1