Negative-stain electron microscopy of inside-out FtsZ rings reconstituted on artificial membrane tubules show ribbons of protofilaments.

Milam, Sara L

Osawa, Masaki

Erickson, Harold P

2018-04-01T14:53:23Z

2018-04-01T14:53:23Z

2012-07

2018-04-01T14:53:22Z

FtsZ, the primary cytoskeletal element of the Z ring, which constricts to divide bacteria, assembles into short, one-stranded filaments in vitro. These must be further assembled to make the Z ring in bacteria. Conventional electron microscopy (EM) has failed to image the Z ring or resolve its substructure. Here we describe a procedure that enabled us to image reconstructed, inside-out FtsZ rings by negative-stain EM, revealing the arrangement of filaments. We took advantage of a unique lipid that spontaneously forms 500 nm diameter tubules in solution. We optimized conditions for Z-ring assembly with fluorescence light microscopy and then prepared specimens for negative-stain EM. Reconstituted FtsZ rings, encircling the tubules, were clearly resolved. The rings appeared as ribbons of filaments packed side by side with virtually no space between neighboring filaments. The rings were separated by variable expanses of empty tubule as seen by light microscopy or EM. The width varied considerably from one ring to another, but each ring maintained a constant width around its circumference. The inside-out FtsZ rings moved back and forth along the tubules and exchanged subunits with solution, similarly to Z rings reconstituted outside or inside tubular liposomes. FtsZ from Escherichia coli and Mycobacterium tuberculosis assembled rings of similar structure, suggesting a universal structure across bacterial species. Previous models for the Z ring in bacteria have favored a structure of widely scattered filaments that are not in contact. The ribbon structure that we discovered here for reconstituted inside-out FtsZ rings provides what to our knowledge is new evidence that the Z ring in bacteria may involve lateral association of protofilaments.

S0006-3495(12)00615-7

0006-3495

1542-0086

https://hdl.handle.net/10161/16457

eng

Elsevier BV

Biophysical journal

10.1016/j.bpj.2012.05.035

Escherichia coli

Mycobacterium tuberculosis

Lipid Bilayers

Bacterial Proteins

Cytoskeletal Proteins

Liposomes

Microscopy, Electron

Microscopy, Fluorescence

Fluorescence Recovery After Photobleaching

Negative Staining

Amino Acid Motifs

Molecular Conformation

Negative-stain electron microscopy of inside-out FtsZ rings reconstituted on artificial membrane tubules show ribbons of protofilaments.

Journal article

Erickson, Harold P|0000-0002-9104-8987

1

School of Medicine

Duke

Duke Cancer Institute

Institutes and Centers

Biochemistry

Basic Science Departments

Cell Biology

Published

103