Abstract:
:Fluorescent proteins are widely used as fusion tags to detect protein expression in vivo. To become fluorescent, these proteins must undergo chromophore maturation, a slow process with a half-time of 5 to >30 min that causes delays in real-time detection of protein expression. Here, we engineer a genetically encoded fluorescent biosensor to enable detection of protein expression within seconds in live bacteria. This sensor for transiently expressed proteins (STEP) is based on a fully matured but dim green fluorescent protein in which pre-existing fluorescence increases 11-fold in vivo following the specific and rapid binding of a protein tag (Kd 120 nM, kon 1.7 × 105 M-1 s-1). In live E. coli cells, our STEP biosensor enables detection of protein expression twice as fast as the use of standard fluorescent protein fusions. Our biosensor opens the door to the real-time study of short timescale processes in live cells with high spatiotemporal resolution.
journal_name
ACS Synth Bioljournal_title
ACS synthetic biologyauthors
Eason MG,Pandelieva AT,Mayer MM,Khan ST,Garcia HG,Chica RAdoi
10.1021/acssynbio.0c00407subject
Has Abstractpub_date
2020-11-20 00:00:00pages
2955-2963issue
11issn
2161-5063journal_volume
9pub_type
杂志文章abstract::Genome engineering technologies now enable precise manipulation of organism genotype, but can be limited in scalability by their design requirements. Here we describe Merlin ( http://merlincad.org ), an open-source web-based tool to assist biologists in designing experiments using multiplex automated genome engineerin...
journal_title:ACS synthetic biology
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journal_title:ACS synthetic biology
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journal_title:ACS synthetic biology
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journal_title:ACS synthetic biology
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