4RZS

Lac repressor engineered to bind sucralose, unliganded tetramer

Summary for 4RZS

• Entry DOI: 10.2210/pdb4rzs/pdb
• Descriptor: Lac repressor, GLYCEROL (3 entities in total)
• Functional Keywords: sucralose, laci, lac repressor, lactose operon repressor, protein design, allostery, escherichia coli, transcription
• Biological source: Escherichia coli
• Total number of polymer chains: 4
• Total formula weight: 163902.33

Authors

Arbing, M.A.,Cascio, D.,Kosuri, S.,Church, G.M. (deposition date: 2014-12-24, release date: 2015-12-23, Last modification date: 2024-02-28)

Primary citation

Taylor, N.D.,Garruss, A.S.,Moretti, R.,Chan, S.,Arbing, M.A.,Cascio, D.,Rogers, J.K.,Isaacs, F.J.,Kosuri, S.,Baker, D.,Fields, S.,Church, G.M.,Raman, S.
Engineering an allosteric transcription factor to respond to new ligands.
Nat.Methods, 13:177-183, 2016

PubMed Abstract: Genetic regulatory proteins inducible by small molecules are useful synthetic biology tools as sensors and switches. Bacterial allosteric transcription factors (aTFs) are a major class of regulatory proteins, but few aTFs have been redesigned to respond to new effectors beyond natural aTF-inducer pairs. Altering inducer specificity in these proteins is difficult because substitutions that affect inducer binding may also disrupt allostery. We engineered an aTF, the Escherichia coli lac repressor, LacI, to respond to one of four new inducer molecules: fucose, gentiobiose, lactitol and sucralose. Using computational protein design, single-residue saturation mutagenesis or random mutagenesis, along with multiplex assembly, we identified new variants comparable in specificity and induction to wild-type LacI with its inducer, isopropyl β-D-1-thiogalactopyranoside (IPTG). The ability to create designer aTFs will enable applications including dynamic control of cell metabolism, cell biology and synthetic gene circuits.

PubMed: 26689263
DOI: 10.1038/nmeth.3696

Experimental method

X-RAY DIFFRACTION (2.71 Å)