Probing for Binding Regions of the FtsZ Protein Surface through Site-Directed Insertions: Discovery of Fully Functional FtsZ-Fluorescent Proteins.
Abstract
FtsZ, a bacterial tubulin homologue, is a cytoskeletal protein that assembles into
protofilaments that are one subunit thick. These protofilaments assemble further to
form a "Z ring" at the center of prokaryotic cells. The Z ring generates a constriction
force on the inner membrane and also serves as a scaffold to recruit cell wall remodeling
proteins for complete cell division in vivo One model of the Z ring proposes that
protofilaments associate via lateral bonds to form ribbons; however, lateral bonds
are still only hypothetical. To explore potential lateral bonding sites, we probed
the surface of Escherichia coli FtsZ by inserting either small peptides or whole fluorescent
proteins (FPs). Among the four lateral surfaces on FtsZ protofilaments, we obtained
inserts on the front and back surfaces that were functional for cell division. We
concluded that these faces are not sites of essential interactions. Inserts at two
sites, G124 and R174, located on the left and right surfaces, completely blocked function,
and these sites were identified as possible sites for essential lateral interactions.
However, the insert at R174 did not interfere with association of protofilaments into
sheets and bundles in vitro Another goal was to find a location within FtsZ that supported
insertion of FP reporter proteins while allowing the FtsZ-FPs to function as the sole
source of FtsZ. We discovered one internal site, G55-Q56, where several different
FPs could be inserted without impairing function. These FtsZ-FPs may provide advances
for imaging Z-ring structure by superresolution techniques. IMPORTANCE: One model
for the Z-ring structure proposes that protofilaments are assembled into ribbons by
lateral bonds between FtsZ subunits. Our study excluded the involvement of the front
and back faces of the protofilament in essential interactions in vivo but pointed
to two potential lateral bond sites, on the right and left sides. We also identified
an FtsZ loop where various fluorescent proteins could be inserted without blocking
function; these FtsZ-FPs functioned as the sole source of FtsZ. This advance provides
improved tools for all fluorescence imaging of the Z ring and may be especially important
for superresolution imaging.
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https://hdl.handle.net/10161/13914Published Version (Please cite this version)
10.1128/JB.00553-16Publication Info
Moore, Desmond A; Whatley, Zakiya N; Joshi, Chandra P; Osawa, Masaki; & Erickson,
Harold P (2017). Probing for Binding Regions of the FtsZ Protein Surface through Site-Directed Insertions:
Discovery of Fully Functional FtsZ-Fluorescent Proteins. J Bacteriol, 199(1). 10.1128/JB.00553-16. Retrieved from https://hdl.handle.net/10161/13914.This is constructed from limited available data and may be imprecise. To cite this
article, please review & use the official citation provided by the journal.
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Show full item recordScholars@Duke
Harold Paul Erickson
James B. Duke Distinguished Professor Emeritus
Recent research has been on cytoskeleton (eukaryotes and bacteria); a skirmish to
debunk the irisin story; a reinterpretation of proposed multivalent binders of the
coronavirus spike protein. I have also published an ebook on "Principles of Protein-Protein
Association" suitable for a course module or individual learning.
Masaki Osawa
Assistant Research Professor of Cell Biology
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