Membrane topography of ColE1 gene products: the immunity protein.
The topography of the colicin E1 immunity (Imm) protein was determined from the positions of TnphoA and complementary lacZ fusions relative to the three long hydrophobic segments of the protein and site-directed substitution of charged for nonpolar residues in the proposed membrane-spanning segments. Inactivation of the Imm protein function required substitution and insertion of two such charges. It was concluded that the 113-residue colicin E1 Imm protein folds in the membrane as three trans-membrane alpha-helices, with the NH2 and COOH termini on the cytoplasmic and periplasmic sides of the membrane, respectively. The approximate spans of the three helices are Asn-9 to Ser-28, Ile-43 to Phe-62, and Leu-84 to Leu-104. An extrinsic highly charged segment, Lys-66 to Lys-74, containing seven charges in nine residues, extends into the cytoplasmic domain. The specificity of the colicin E1 Imm protein for interaction with the translocation apparatus and the colicin E1 ion channel is proposed to reside in its peripheral segments exposed on the surface of the inner membrane. These regions include the highly charged segment Lys-66 to Lys-83 (loop 2) and the short (approximately eight-residue) NH2 terminus on the cytoplasmic side, and Glu-29 to Val-44 (loop 1) and the COOH-terminal segment Gly-105 to Asn-113 on the periplasmic side.
Song
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Department of Biological Sciences, Purdue University, West Lafayette, Indiana 47907.
Cramer
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D000469:Alkaline Phosphatase / Q000378:metabolism; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000737:chemistry; D001692:Biological Transport; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure*; D003001:Cloning, Molecular; D003087:Colicins / Q000037:antagonists & inhibitors*; D007473:Ion Channels; D008954:Models, Biological; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D016297:Mutagenesis, Site-Directed; D010957:Plasmids; D012689:Sequence Homology, Nucleic Acid; D001616:beta-Galactosidase / Q000378:metabolism
D001426:Bacterial Proteins; D003087:Colicins; D007473:Ion Channels; C007104:colicin immunity proteins; D000469:Alkaline Phosphatase; D001616:beta-Galactosidase