Open AccessCharacterization of the pColV‐K30 encoded cloacin DF13/aerobactin outer membrane receptor protein of Escherichia coli ; isolation and purification of the protein and analysis of its nucleotide sequence and primary structure
Author(s) -
Krone Willy J.A.,
Stegehuis Freek,
Koningstein Gregory,
Doorn Carla,
Roosendaal Bert,
Graaf Frits K.,
Oudega Bauke
Publication year - 1985
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.1985.tb01583.x
Subject(s) - aerobactin , escherichia coli , camp receptor protein , colicin , bacterial outer membrane , biology , microbiology and biotechnology , affinity chromatography , biochemistry , fusion protein , peptide sequence , chemistry , enterobacteriaceae , recombinant dna , gene , enzyme , gene expression , promoter
The cloacin DF13/aerobactin receptor protein from Escherichia coli (pFS8) and from Klebsiella edwardsii were isolated by repeated Triton X‐100 extractions and purified by affinity chromatography. Both receptor proteins ran as a single protein band on SDS‐PAGE. Their apparent M r values were 74 000 and 76 000, respectively. The binding constants of the purified receptor proteins from E. coli (pFS8) and K. edwardsii and cloacin DF13 were determined. Values of 2.0 × 10 8 M −1 and 1.0 × 10 9 M −1 , respectively, were found. The nucleotide sequence of the pColV‐K30 gene, contained on pFS8 and encoding the cloacin DF13/aerobactin receptor protein, was determined and the primary structure of the protein as well as its secondary structure were deduced. The results revealed that the pColV‐K30‐specified receptor protein might be synthesized as a precursor, with a signal sequence of 25 amino acid residues. The mature protein has an M r of 77 345.