TY  - JOUR
A1  - Brechun, Katherine Emily
A1  - Arndt, Katja Maren
A1  - Woolley, G. Andrew
T1  - Selection of protein-protein interactions of desired affinities with a bandpass circuit
JF  - Journal of molecular biology : JMB
N2  - We have developed a genetic circuit in Escherichia coli that can be used to select for protein-protein interactions of different strengths by changing antibiotic concentrations in the media. The genetic circuit links protein-protein interaction strength to beta-lactamase activity while simultaneously imposing tuneable positive and negative selection pressure for beta-lactamase activity. Cells only survive if they express interacting proteins with affinities that fall within set high- and low-pass thresholds; i.e. the circuit therefore acts as a bandpass filter for protein-protein interactions. We show that the circuit can be used to recover protein-protein interactions of desired affinity from a mixed population with a range of affinities. The circuit can also be used to select for inhibitors of protein-protein interactions of defined strength. (C) 2018 Elsevier Ltd. All rights reserved.
KW  - synthetic biology
KW  - genetic circuit
KW  - biological engineering
KW  - protein-protein interactions
KW  - twin-arginine translocation
KW  - selection system
Y1  - 2018
U6  - https://doi.org/10.1016/j.jmb.2018.11.011
SN  - 0022-2836
SN  - 1089-8638
VL  - 431
IS  - 2
SP  - 391
EP  - 400
PB  - Elsevier
CY  - London
ER  - 
TY  - JOUR
A1  - Brechun, Katherine E.
A1  - Arndt, Katja Maren
A1  - Woolley, G. Andrew
T1  - Strategies for the photo-control of endogenous protein activity
JF  - Current opinion in structural biology : review of all advances ; evaluation of key references ; comprehensive listing of papers
Y1  - 2017
U6  - https://doi.org/10.1016/j.sbi.2016.11.014
SN  - 0959-440X
SN  - 1879-033X
VL  - 45
SP  - 53
EP  - 58
PB  - Elsevier
CY  - London
ER  - 
TY  - JOUR
A1  - Brechun, Katherine E.
A1  - Zhen, Danlin
A1  - Jaikaran, Anna
A1  - Borisenko, Vitali
A1  - Kumauchi, Masato
A1  - Hoff, Wouter D.
A1  - Arndt, Katja Maren
A1  - Woolley, Andrew G
T1  - Detection of Incorporation of p-Coumaric Acid into Photoactive Yellow Protein Variants in Vivo
JF  - Biochemistry
N2  - We report the design and characterization of photoactive yellow protein (PYP)-blue fluorescent protein (mTagBFP) fusion constructs that permit the direct assay of reconstitution and function of the PYP domain. These constructs allow for in vivo testing of co-expression systems for enzymatic production of the p-coumaric acid-based PYP chromophore, via the action of tyrosine ammonia lyase and p-coumaroyl-CoA ligase (pCL or 4CL). We find that different 4CL enzymes can function to reconstitute PYP, including 4CL from Arabidopsis thaliana that can produce similar to 100% holo-PYP protein under optimal conditions. mTagBFP fusion constructs additionally enable rapid analysis of effects of mutations on PYP photocycles. We use this mTagBFP fusion strategy to demonstrate in vivo reconstitution of several PYP-based optogenetic tools in Escherichia coli via a biosynthesized chromophore, an important step for the use of these optogenetic tools in vivo in diverse hosts.
Y1  - 2019
U6  - https://doi.org/10.1021/acs.biochem.9b00279
SN  - 0006-2960
VL  - 58
IS  - 23
SP  - 2682
EP  - 2694
PB  - American Chemical Society
CY  - Washington
ER  - 
TY  - JOUR
A1  - Mazumder, Mostafizur
A1  - Brechun, Katherine E.
A1  - Kim, Yongjoo B.
A1  - Hoffmann, Stefan A.
A1  - Chen, Yih Yang
A1  - Keiski, Carrie-Lynn
A1  - Arndt, Katja Maren
A1  - McMillen, David R.
A1  - Woolley, G. Andrew
T1  - An Escherichia coli system for evolving improved light-controlled DNA-binding proteins
JF  - Protein engineering design & selection
N2  - Light-switchable proteins offer numerous opportunities as tools for manipulating biological systems with exceptional degrees of spatiotemporal control. Most designed light-switchable proteins currently in use have not been optimised using the randomisation and selection/screening approaches that are widely used in other areas of protein engineering. Here we report an approach for screening light-switchable DNA-binding proteins that relies on light-dependent repression of the transcription of a fluorescent reporter. We demonstrate that the method can be used to recover a known light-switchable DNA-binding protein from a random library.
KW  - directed evolution
KW  - fluorescent reporter
KW  - optogenetics
Y1  - 2015
U6  - https://doi.org/10.1093/protein/gzv033
SN  - 1741-0126
SN  - 1741-0134
VL  - 28
IS  - 9
SP  - 293
EP  - 302
PB  - Oxford Univ. Press
CY  - Oxford
ER  -