The DNA target of the trp repressor.
Unexpected features seen by high resolution X-ray crystallography at the interface of the trp repressor and the 'traditional' trp operator provoked the claim that the DNA fragment used in the crystal structure is not the true operator, and therefore that the crystal structure of the trp repressor-operator complex does not portray a specific interaction. An alternative sequence was proposed mainly on the basis of mutational studies and gel retardation analysis of short target duplexes (Staacke et al., 1990a,b). We have reexamined the sequence consensus in trpR-repressible promoters and analyzed the mutagenesis experiments of others including Staacke et al. (1990a) and found them fully consistent with the interactions of the traditional operator sequence seen in the crystal structure, and stereochemically inconsistent with the above referenced alternative model. Moreover, an in vitro trp repressor-DNA binding analysis, employing both novel DNA constructs devised to avoid previously encountered artifacts as well as full-length promoter sequences, indicates that the traditional operator used in the crystal structure is the preferred target of the trp repressor.
Haran
T E
TE
Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT 06511.
Joachimiak
A
A
Sigler
P B
PB
D001426:Bacterial Proteins / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009690:Nucleic Acid Conformation; D009838:Oligodeoxyribonucleotides / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012097:Repressor Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014961:X-Ray Diffraction; D029968:Escherichia coli Proteins
D001426:Bacterial Proteins; D004247:DNA; D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins; D012097:Repressor Proteins; C027190:TRPR protein, E coli; D029968:Escherichia coli Proteins