A two-hybrid system based on chimeric operator recognition for studying protein homo/heterodimerization in Escherichia coli.
The development of a convenient and promising alternative to the various two-hybrid methods that are used to study protein-protein interactions is described. In this system, a lambdoid chimeric operator is recognized by a hybrid repressor formed by two chimeric monomers whose C-terminal domains are composed of heterologous proteins (or protein domains). Only if these proteins efficiently dimerize in vivo is a functional repressor formed able to bind the chimeric operator and shut off the synthesis of a downstream reporter gene. This new approach was tested with several interacting proteins ranging in size from less than 100 to more than 800 amino acids and, to date, no size or topology limit has been detected.
Di Lallo
G
G
Dipartimento di Biologia, Università 'Tor Vergata', via della Ricerca Scientifica, 00133 La Romanina (Roma), Italy1.
Castagnoli
L
L
Dipartimento di Biologia, Università 'Tor Vergata', via della Ricerca Scientifica, 00133 La Romanina (Roma), Italy1.
Ghelardini
P
P
Centro Acidi Nucleici del CNR, Roma, Italy2.
Paolozzi
L
L
Dipartimento di Biologia, Università 'Tor Vergata', via della Ricerca Scientifica, 00133 La Romanina (Roma), Italy1.
D001435:Bacteriophages / Q000235:genetics; D019281:Dimerization; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D017930:Genes, Reporter; D009875:Operator Regions, Genetic; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000235:genetics* / Q000378:metabolism; D012097:Repressor Proteins / Q000378:metabolism; D015203:Reproducibility of Results; D020798:Two-Hybrid System Techniques; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D001616:beta-Galactosidase