Bacterial infection of host cells: Pathogens of the type Salmonella typhimurium (orange) establish contact to a human host cell (blue). Image: Christian Goosmann, Diane Schad, Rashmi Gupta and Michael Kolbe

The
plague, bacterial dysentery, and cholera have one thing in common:
These dangerous diseases are caused by bacteria which infect their host
using a sophisticated injection apparatus. Through needle-like
structures, they release molecular agents into their host cell, thereby
evading the immune response. Researchers at the Max Planck Institute for
Biophysical Chemistry in Göttingen in cooperation with colleagues at
the Max Planck Institute for Infection Biology in Berlin and the
University of Washington in Seattle (USA) have now elucidated the
structure of such a needle at atomic resolution. Their findings might
contribute to drug tailoring and the development of strategies which
specifically prevent the infection process.

Hundreds
of tiny hollow needles sticking out of the bacterial membrane—it is a
treacherous tool that makes pathogens causing plague or cholera so
dangerous. Together with a base, embedded in the membrane, these
miniature syringes constitute the so-called type III secretion system –
an injection apparatus through which the pathogens introduce molecular
agents into their host cell. There, these substances manipulate
essential metabolic processes and disable the immune defense of the
infected cells. The consequences are fatal as the pathogens can now
spread within the organism without hindrance. To date, traditional
antibiotics are prescribed to fight the infection. However, as some
bacterial strains succeed in developing resistances, researchers
worldwide seek to discover more specific drugs.

The
exact structure of the 60 to 80 nm long and about 8 nm wide
needles has so far been unknown. Classical methods such as X-ray
crystallography or electron microscopy failed or yielded wrong model
structures. Not crystallizable and insoluble, the needle resisted all
attempts to decode its atomic structure. Therefore Adam Lange and Stefan
Becker at the Max Planck Institute  for Biophysical Chemistry together
with a team of physicists, biologists and chemists chose a completely
novel approach. In cooperation with David Baker at the University of
Washington, and Michael Kolbe at the Max Planck Institute for Infection
Biology, the scientists successfully combined the production of the
needle in the laboratory with solid-state NMR spectroscopy, electron
microscopy, and computer modelling. The researchers deciphered the
structure of the needle atom by atom and visualized its molecular
architecture for the first time in the angstrom range, a resolution of
less than a tenth of a millionth of a millimeter.

This
required progresses in several fields. “We have made big steps forward
concerning sample production as well as solid-state NMR spectroscopy,”
says Adam Lange. “Finally, we were also able to use one of the presently
most powerful solid-state NMR spectrometers in Christian Griesinger’s
NMR-based Structural Biology Department at our Institute.”

With 20 tesla, the magnetic field of this 850 MHz spectrometer is about 400,000 times as strong as that of the earth.

“We
were surprised to see how the needles are constructed,” says Lange. As
expected, the needles of pathogens causing diseases as diverse as food
poisoning, bacterial dysentery, or the plague show striking
similarities. However, in contrast to prevailing assumptions, the
similarities are found in the inner part of the needles whereas the
surface is astonishingly variable.

Syringes isolated from Shigella flexneri. Adding soluble needle protein leads to a spontaneous elongation of some needles. The bar corresponds to 100 nm. Image: MPI for Biophysical Chemistry, Christian Goosmann, Michael Kolbe

 

According
to the scientist, this variability might be a strategy of the bacteria
to evade immune recognition by the host. Changes on the surface of the
needle make it difficult for the host’s immune system to recognize the
pathogen.

The
scientists Lange, Kolbe, Becker, and their Max Planck colleagues
Christian Griesinger und Arturo Zychlinsky, have focused on the
bacterial injection apparatus for several years. Together with the
Federal Institute for Materials Research and Testing they already showed
in 2010 how bacteria assemble their miniature syringes. The discovery
of their structure in atomic detail not only enables researchers to
 gain new insights into how these pathogens outwit their host cells, it
also offers the prospect to block the syringe assembly and the delivery
of the bacterial factors using tailored molecules. Such substances,
referred to as anti-infectives, could act more specifically and much
earlier during infection than traditional antibiotics.

“Thanks
to our new technique, we can produce large amounts of needles in the
lab. Our aim is now to develop a high-throughput method. This will allow
us to search for new agents that prevent the formation of the needle,”
explains Stefan Becker.

Atomic Model of the Type III Secretion System Needle

Nature advance online publication: DOI: 10.1038/nature11079

Source: Max Planck Institute