Architecture and substrate-recognition assembly of the Type IV coupling protein complex of L. pneumophila
The type IV coupling protein (T4CP) complex of Legionella pneumophila is termed a “gatekeeper” for secreting toxic effector proteins into human macrophage cells. The complex plays a key role in the pathogenesis of L. pneumophila.
However, the effector translocation mechanism of the T4CP complex has remained poorly understood. In this study, a biological research team at KAIST recently determined the architecture of the T4CP complex composed of the six proteins abbreviated as: DotL, DotN, DotM, IcmS, IcmW, LvgA. Furthermore, the group identified the substrate-recognition assembly of the T4CP complex.
These results provide important insights into the underlying substrate recruitment and processing mechanism.
Many bacteria translocate virulent effector proteins into a host cell. To establish a multiplication niche, the type IV secretion system (T4SS) is studied as it serves as a special translocation machine in various bacteria.
This system is composed mainly of two components: a “transenvelope secretion conduit” and a “type IV coupling protein (T4CP).” To deliver appropriate substrates, T4SS should distinguish bacterial housekeeping proteins and translocating substrates.
The type IV coupling protein (T4CP) is a hexameric ATPase that selectively recruits translocating substrates and conveys them to the secretion conduit. However, the translocating mechanism of the T4CP has until this time been poorly understood.
Recent results by researchers at KAIST (Department of Biological Sciences) have provided important clues to this enigmatic mechanism. The researchers have studied the T4CP of Legionella pneumophila which causes Legionnaires’ disease.
L. pneumophila secretes ~300 virulent effector proteins into human macrophage cells using T4SS. The secreted effector proteins disturb the immune defense mechanisms to survive and multiply in the infected cell.
To elucidate the effector translocation mechanism, they determined the structure of a T4CP subcomplex. T4CP is composed of three parts; a transmembrane domain, an ATPase domain and a C-terminal extension (CTE).
Three accessory proteins (DotN, IcmS, IcmW) forms a complex with T4CP by interacting with CTE. Moreover, the new binding partner, LvgA, was identified as another CTE-binding component.
Using techniques such as X-ray crystallography, SAXS and ALEX-FRET methods, the scientists determined the structure of the CTE complex consisting of five proteins, DotL, DotN, IcmS, IcmW, LvgA. (figure shown below)
Furthermore, they demonstrated the interaction of the CTE complex and effector proteins. Of the five accessory components, LvgA played a key role in their interaction.
Effector proteins could not bind to the CTE complex lacking LvgA, strongly suggesting that LvgA plays a crucial role in recognizing effector proteins.
The researchers also built a model of the T4CP holocomplex structure. By using homology modeling, the hexameric ATPase domain was built and the CTE complex was anchored to the ATPase domain.
The model is an elongated bell-shaped architecture composed of the membrane-proximal ATPase hexamer and the membrane-distal C-terminal assemblies of DotL (figure shown below).
Based on the architecture of the T4CP holocomplex, the translocation mechanism can now be inferred. Effectors are firstly recruited to bottom lim of the T4CP complex.
It would be processed to a translocatable formation in the T4CP chamber. After processing, it would be translocated via a narrow channel of the ATPase hexamer. A detailed mechanism is needed to reach further into this research topic.
This study is first structural approach taken regarding the T4CP of L. pneumophila. This research was published recently online in Nature Microbiology (July, 2017); the report was entitled, “Architecture of the type IV coupling protein complex of Legionella pneumophila.”
Mi-Jeong Kwak, J. Dongun Kim, Hyunmin Kim, Cheolhee Kim, James W. Bowman, Seonghoon Kim, Keehyoung Joo, Jooyoung Lee, Kyeong Sik Jin, Yeon-Gil Kim, Nam Ki Lee, Jae U. Jung and Byung-Ha Oh.
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* lab webpage : http://struct.kaist.ac.kr/