Visualization of Spirochetes by Labeling Membrane Proteins With Fluorescent Biarsenical Dyes

Result code in IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F60077344%3A_____%2F19%3A00519670" target="_blank" >RIV/60077344:_____/19:00519670 - isvavai.cz</a>

Alternative codes found

RIV/60076658:12310/19:43899625

Result on the web

<a href="https://www.frontiersin.org/articles/10.3389/fcimb.2019.00287/full" target="_blank" >https://www.frontiersin.org/articles/10.3389/fcimb.2019.00287/full</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.3389/fcimb.2019.00287" target="_blank" >10.3389/fcimb.2019.00287</a>

Result language

angličtina

Original language name

Visualization of Spirochetes by Labeling Membrane Proteins With Fluorescent Biarsenical Dyes

Original language description

Numerous methods exist for fluorescently labeling proteins either as direct fusion proteins (GFP, RFP, YFP, etc.-attached to the protein of interest) or utilizing accessory proteins to produce fluorescence (SNAP-tag, CLIP-tag), but the significant increase in size that these accompanying proteins add may hinder or impede proper protein folding, cellular localization, or oligomerization. Fluorescently labeling proteins with biarsenical dyes, like FlAsH, circumvents this issue by using a short 6-amino acid tetracysteine motif that binds the membrane-permeable dye and allows visualization of living cells. Here, we report the successful adaptation of FlAsH dye for live-cell imaging of two genera of spirochetes, Leptospira and Borrelia, by labeling inner or outer membrane proteins tagged with tetracysteine motifs. Visualization of labeled spirochetes was possible by fluorescence microscopy and flow cytometry. A subsequent increase in fluorescent signal intensity, including prolonged detection, was achieved by concatenating two copies of the 6-amino acid motif. Overall, we demonstrate several positive attributes of the biarsenical dye system in that the technique is broadly applicable across spirochete genera, the tetracysteine motif is stably retained and does not interfere with protein function throughout the B. burgdorferi infectious cycle, and the membrane-permeable nature of the dyes permits fluorescent detection of proteins in different cellular locations without the need for fixation or permeabilization. Using this method, new avenues of investigation into spirochete morphology and motility, previously inaccessible with large fluorescent proteins, can now be explored.

Czech name

—

Czech description

—

Type

J_imp - Article in a specialist periodical, which is included in the Web of Science database

CEP classification

—

OECD FORD branch

20201 - Electrical and electronic engineering

Project

<a href="/en/project/TE01020118" target="_blank" >TE01020118: Electron microscopy</a><br>

Continuities

I - Institucionalni podpora na dlouhodoby koncepcni rozvoj vyzkumne organizace

Publication year

2019

Confidentiality

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Name of the periodical

Frontiers in Cellular and Infection Microbiology

ISSN

2235-2988

e-ISSN

—

Volume of the periodical

9

Issue of the periodical within the volume

AUG 20 2019

Country of publishing house

CH - SWITZERLAND

Number of pages

14

Pages from-to

287

UT code for WoS article

000481779200001

EID of the result in the Scopus database

2-s2.0-85071773412