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Name (Title):
Tomohiko YAMAZAKI (MANA Scientist)
Affiliation:
International Center for Materials Nanoarchitectonics
(MANA), NIMS
Address:
1-1 Namiki, Tsukuba. Ibaraki, 305-0044, Japan
1-2
Email: YAMAZAKI.Tomohiko@nims.go.jp
Home Page: http://www.nims.go.jp/bmc/group/medical/index_e.html
Presentation Title:
Building a novel transcriptional regulator as a universal switch platform
Abstract:
Transcriptional regulators activated by the target molecule interact with the promoter triggering
production of the functional proteins. Utilizing transcriptional regulators, biosensors, transgenic
expression systems, and signal amplification cascades can be constructed that detect compounds as
varied as those related to quorum sensing. Engineering of target molecule–binding site of
transcriptional regulator to enhance sensitivity and specificity to target molecules have been
reported. We have also proposed a novel design and construction method for engineered
transcriptional regulators.
Bacterial substrate binding proteins (bSBPs) have been focused as novel probes to biosensing
and biosensors. bSBPs have been identified for a wide variety of ligands with high affinity (Kd =
approx. 10 -6 M). Most bSBPs have a similar structural consisting of two domains linked by a hinge
region and undergo a large conformational change by ligand binding. The developments of bSBPbased
sensing systems are recently reported by screening of novel bSBPs for desired target or by
genetic engineering to improve its binding properties, etc. bSBPs have thus great possibility to be
a universal probe which can be applied to sensing systems for variety of ligands.
Here we report our novel approach in the design and construction of engineered transcriptional
regulators based on engineered bSBPs as target molecule recognition elements. We designed
bSBP-based regulators, bSBP-LTR chimeric
proteins (SLCP) combined one of bSBPs with a
bSBP-like regulator, LacI-like repressor proteins
(LTR), which consists of DNA-binding domain
and substrate binding domain similar to bSBPs A
chimeric protein of well analyzed bSBP,
galactose-/glucose -binding protein (GBP), and
LacI from Eschelichia coli was at first selected as
a model of bSBP-based regulators and constructed
based on 3-D structural models. We also show the
characterization of the constructed GBP-LacI
chimeric protein as a transcriptional regulator in
vitro and in mammalian cells.
Poster Session PB-5
Figure Construction of novel transcriptional
regulators
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