Biomaterial Database

Operator search by mutant Lac repressors. 1998

A Barker, and R Fickert, and S Oehler, and B Müller-hill

Institut für Genetik der Universität zu Köln, Köln, Weyertal 121, D-50931, Germany.

The Escherichia coli Lac and Gal repressors are two members of a large family of bacterial repressor proteins that share significant sequence and structural homology. Efficient repression by all family members requires specific binding to a site or sites close to the transcriptional start of the genes regulated. Both LacR and GalR have to bind to at least two sites for efficient repression, yet they differ in one important respect: LacR is a homotetramer whereas GalR is a homodimer. In an attempt to understand this difference, we studied the operator binding activity of a LacR variant that has the DNA-binding specificity of GalR (LacR-V17A18). A tetrameric version of this protein shows a 30-fold decrease in association rate to operator located on a long (lambda) DNA molecule, in comparison to wild-type LacR, while a dimeric version of this protein shows an unaltered association rate in comparison to dimeric LacR. This reduction in association rate correlates with a broadened DNA-binding specificity for base-pairs 4 and 5 of the operator: examination of an additional LacR variant with an even broader DNA-binding specificity indicates that a tetrameric version also shows a 30-fold decrease in association rate in comparison to wild-type LacR, while a dimeric version again shows an unaltered association rate in comparison to dimeric LacR. This difference in association rate in vitro correlates with whether a tetrameric or dimeric variant of LacR of a given DNA-binding specificity will repress lacZ under control of a single operator more efficiently in vivo. We therefore propose that the formation of stable homotetramers becomes a distinct disadvantage unless a high degree of DNA-binding specificity is also present, and demonstrate that this in indeed the case for GalR-mediated repression of the gal operon. This functional constraint seems to have influenced the evolution of the LacI-GalR family of repressors, most of which have a relatively broad specificity of DNA-binding and most of which form only stable homodimers.

UI	MeSH Term	Description	Entries
D007700	Kinetics	The rate dynamics in chemical or physical systems.
D008958	Models, Molecular	Models used experimentally or theoretically to study molecular shape, electronic properties, or interactions; includes analogous molecules, computer-generated graphics, and mechanical structures.	Molecular Models,Model, Molecular,Molecular Model
D009876	Operon	In bacteria, a group of metabolically related genes, with a common promoter, whose transcription into a single polycistronic MESSENGER RNA is under the control of an OPERATOR REGION.	Operons
D010582	Bacteriophage lambda	A temperate inducible phage and type species of the genus lambda-like viruses, in the family SIPHOVIRIDAE. Its natural host is E. coli K12. Its VIRION contains linear double-stranded DNA with single-stranded 12-base 5' sticky ends. The DNA circularizes on infection.	Coliphage lambda,Enterobacteria phage lambda,Phage lambda,lambda Phage
D010957	Plasmids	Extrachromosomal, usually CIRCULAR DNA molecules that are self-replicating and transferable from one organism to another. They are found in a variety of bacterial, archaeal, fungal, algal, and plant species. They are used in GENETIC ENGINEERING as CLONING VECTORS.	Episomes,Episome,Plasmid
D012097	Repressor Proteins	Proteins which maintain the transcriptional quiescence of specific GENES or OPERONS. Classical repressor proteins are DNA-binding proteins that are normally bound to the OPERATOR REGION of an operon, or the ENHANCER SEQUENCES of a gene until a signal occurs that causes their release.	Repressor Molecules,Transcriptional Silencing Factors,Proteins, Repressor,Silencing Factors, Transcriptional
D004269	DNA, Bacterial	Deoxyribonucleic acid that makes up the genetic material of bacteria.	Bacterial DNA
D004926	Escherichia coli	A species of gram-negative, facultatively anaerobic, rod-shaped bacteria (GRAM-NEGATIVE FACULTATIVELY ANAEROBIC RODS) commonly found in the lower part of the intestine of warm-blooded animals. It is usually nonpathogenic, but some strains are known to produce DIARRHEA and pyogenic infections. Pathogenic strains (virotypes) are classified by their specific pathogenic mechanisms such as toxins (ENTEROTOXIGENIC ESCHERICHIA COLI), etc.	Alkalescens-Dispar Group,Bacillus coli,Bacterium coli,Bacterium coli commune,Diffusely Adherent Escherichia coli,E coli,EAggEC,Enteroaggregative Escherichia coli,Enterococcus coli,Diffusely Adherent E. coli,Enteroaggregative E. coli,Enteroinvasive E. coli,Enteroinvasive Escherichia coli
D005690	Galactose	An aldohexose that occurs naturally in the D-form in lactose, cerebrosides, gangliosides, and mucoproteins. Deficiency of galactosyl-1-phosphate uridyltransferase (GALACTOSE-1-PHOSPHATE URIDYL-TRANSFERASE DEFICIENCY DISEASE) causes an error in galactose metabolism called GALACTOSEMIA, resulting in elevations of galactose in the blood.	D-Galactose,Galactopyranose,Galactopyranoside,D Galactose
D005838	Genotype	The genetic constitution of the individual, comprising the ALLELES present at each GENETIC LOCUS.	Genogroup,Genogroups,Genotypes