Characterization of the nucM gene coding for a nuclease of the phytopathogenic bacteria Erwinia chrysanthemi.
The gene nucM encoding a nuclease was cloned from a genomic library of Erwinia chrysanthemi. The nucM gene was subcloned, and mutagenized by insertion of a uidA-KanR cartridge. This mutation was introduced by recombination into the Erwinia chrysanthemi chromosome. The nucM mutant lost NucM activity when tested on a DNA plate after 24 hours, but still possessed secondary weak nuclease activity. The nucleotide sequence of nucM was determined. It presents a 798 bp open reading frame, coding for a 266-amino-acid protein, with a predicted molecular mass of 29,910 Da. The deduced NucM protein shows 59% sequence identity with the DNase I precursor from Vibrio cholerae. It contains a typical leader sequence. Experiments of cell fractionation showed that NucM is periplasmic in E. chrysanthemi. The transcription start has been determined by S1 mapping. The -10 and -35 regions do not show homology with consensus sequence of the promoters recognized by sigma 70. In fact, the promoter seems to be dependent on the sigma 70, but the first transcription nucleotide is unusually far from the -10 region. nucM seems to be expressed constitutively.
Moulard
M
M
Laboratoire de Génétique Moléculaire des Microorganismes, URA-CNRS 1486, Villeurbanne, France.
Condemine
G
G
Robert-Baudouy
J
J
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics*; D001483:Base Sequence; D002451:Cell Compartmentation; D003001:Cloning, Molecular; D003851:Deoxyribonucleases; D016974:Dickeya chrysanthemi / Q000201:enzymology / Q000235:genetics*; D005966:Glucuronidase / Q000096:biosynthesis / Q000235:genetics; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D011401:Promoter Regions, Genetic / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D012689:Sequence Homology, Nucleic Acid; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D011993:Recombinant Fusion Proteins; D003851:Deoxyribonucleases; C082158:nucM protein, Erwinia chrysanthemi; D005966:Glucuronidase