Dr Fasseli Coulibaly
Senior Research Fellow
ARC Future Fellowship
Phone: +61 3 990 29225
Fax: +61 3 990 29500
Office: Room G29, Building 76, Clayton
Our research aims at understanding the molecular mechanisms underlying the infectious cycles of animal and human viruses using a structural biology approach.
THE STRUCTURAL VIROLOGY LABORATORY
Understanding how viruses assemble and interact with their cellular hosts not only helps fighting off viral pathogens but it also provides a powerful probe to analyse many fundamental processes in biology. The study of poxviruses has contributed to both of these aims. Poxviruses have been a paradigm in vaccination that culminated in the eradication of smallpox by the end of the 1970's. In addition, poxviruses have been model systems because of their complex infectious cycle and their remarkable ability to hijack and neutralize cellular defense mechanisms. In particular, the assembly of vaccinia has been extensively studied by electron microscopy of infected cells and, more recently, by tomography of infectious vaccinia virus particles. Building on this knowledge, our research aims at elucidating the molecular details of poxvirus assembly using X-ray crystallography.
Scaffolding proteins in poxvirus assembly
Contrary to most enveloped viruses, poxviruses do not acquire their internal membrane by budding through cellular compartments. Instead they mature from crescent-shaped precursors whose origin and structure remains controversial. This process results in the formation of non-infectious spherical particles called immature virions or IV.
To investigate the mechanisms underlying this essential maturation step, we focus on a scaffolding protein called D13 in vaccinia virus, the prototype poxvirus. This protein forms a honeycomb scaffold on the surface of IV. Its central role in poxvirus assembly is highlighted by the fact that point mutations of D13 block morphogenesis before the formation of IV. Also the antibiotic rifampicin induces a similar arrest in vaccinia virus morphogenesis that is overcome by point mutations in D13.
Our structure-function studies of the formation of crescents and IV aim at understanding how poxvirus acquire and remodel their membrane, a fundamental process possibly shared by a large group of DNA viruses. This research may also provide new targets for the development of antivirals specifically blocking the assembly of poxviruses.
Spheroids: crystalline armours of poxviruses
Poxvirus are unusual in the virus world because they exhibit several infectious particles differing both by their structures and cellular localisations. Among these infectious forms, spheroids of insect poxviruses are probably the most striking.
Spheroids are crystals forming inside infected cells and visible by light microscopy. They function as a dense matrix that protects virus particles from environmental insults so that they remain viable for years in soil, analogous to bacterial spores. Recently, we have developed methods allowing the structural analysis of such infectious crystals called polyhedra produced by two unrelated viruses, cypovirus and baculovirus. We are now extending these cutting-edge microcrystallography approaches to study spheroids with the aim of unravelling the evolutionary origin of this unique survival strategy and advancing our understanding of in vivo protein crystallization.
In addition, the protective function and the robustness of these crystals has significant applied outcomes, in that they form natural platforms for the development of strong and versatile nanocontainers for example to design novel vaccine vectors.
MicroCubes: natural crystals as antigen carriers
Polyhedra are protein crystals formed in vivo by insect viruses. Their role in the viral cycle is to embed virus particles and protect them in the environment, which means that polyhedra have remarkable robustness and packaging ability. We are interested in exploiting these features to engineer microparticles as a new platform for vaccination (www.monash.edu.au/assets/pdf/industry/microcube-info.pdf)
Structure of birnavirusesBirnaviruses are ubiquitous animal viruses infecting hosts ranging from insects and molluscs to fishes and birds. The prototype virus is the Infectious Bursal Disease Virus (IBDV)
Microcrystallography of viral polyhedra
Our structural analysis of the silkworm cypovirus-1 (CPV-1) polyhedra has allowed the determination of the first atomic structure of viral infectious crystals. These are among the smallest crystals used for solving a protein structure by X-ray crystallography (5-12 micrometers in diameter; data collected at beamline X06SA of the Swiss Light Source). Our results suggest molecular bases for the outstanding stability of polyhedra in the environment as well as for the rapid dissolution at the alkaline pH encountered in the mid-guts of larvae. The structure of CPV-1 polyhedra also provides a platform for the design of protein nanocontainers protecting and delivering valuable cargos such as fluorescent probes or drugs.
Structure of the pili of streptococcus pyogenens
Hair-like appendages on the surface of many gram-negative bacteria, commonly called pili, participate to the process of invasion of target tissues. They constitute important virulence factors and their assembly and function have been well established. In contrast, gram-positive bacteria have long been considered as devoid of pili before the recent identification of pili in a virulence island of Streptococci species. We determined the 2Å structure of the backbone pili subunit of Streptococcus pyogenens and proposed a model for the whole pili assembly. These pili are head-to-tail, covalent polymers of the backbone subunit with sortase-catalysed inter-molecular isopeptide bonds. We also identified two self-generated, intra-molecular isopeptides stabilising each subunit. These bonds allow the pili to resist intense mechanical stress when binding target tissues. A 3-D template search of the PDB revealed that such isopeptide bonds are a common feature of gram-positive bacterial surface proteins that has been overlooked until now.
Hyun JK, Accurso C, Hijnen M, Schult P, Pettikiriarachchi A, Mitra AK* and Coulibaly F* (2011). Membrane remodeling by the double-barrel scaffolding protein of poxvirus. PLoS Pathogens 7:e1002239. * corresponding authors
Gutsche I, Coulibaly F, Voss J, Salmon J, d'Alayer J, Ermonval M, Larquet E, Charneau P, Krey T, Megret F, Guittet E, Rey FA, Flamand M (2011). Secreted dengue virus nonstructural protein NS1 is an atypical barrel-shaped high density lipoprotein. Proc Natl Acad Sci USA 108:8003-8008
Kang H.J., Coulibaly F., Proft T. and Baker E.N. (2011). Crystal structure of Spy0129, a Streptococcus pyogenes class B sortase involved in pilus biogenesis. Plos One 6:e15969.
Law, Lukoyanova, Voskoboinik, Caradoc-Davies, Baran, Dunstone, D’Angelo, Orlova, Coulibaly, Verschoor, Browne, Ciccone, Kuiper, Bird, Trapani, Saibil and Whisstock. (2010) The structural basis for membrane binding and pore formation by lymphocyte perforin. Nature 468:447-51.
Coulibaly F., Chevalier C., Delmas B. and Rey F.A. (2010). Crystal structure of an Aquabirnavirus particle: insights into antigenic diversity and virulence determinism. J. Virol. 84:1792-9.
Ohtsuka Y., Nakai D., Coulibaly F., Chiu E., Metcalf P., Mori H. (2010). Mutations of cypovirus polyhedrin and applications of polyhedra to protein nanocontainers. Nanotech, (CRC Press). Vol 3, pp292-295.
Volkoff A.-N., Jouan V., Urbach S., Samain S., Bergoin M., Wincker P., Demettre E., Cousserans F., Provost B., Coulibaly F., Legeai F., Beliveau C., Cusson M., Gyapay G., Drezen J.-M. (2010) Analysis of Virion Structural Components Reveals Vestiges of the Ancestral Ichnovirus Genome. PLoS Pathogens 6:e1000923 EP
Coulibaly F.*, Chiu E., Ikeda K., Gutmann S., Haebel P.W., Schulze-Briese C., Mori H. and Metcalf, P.* (2009). The atomic structure of baculovirus polyhedra reveals the independent emergence of infectious crystals in DNA and RNA viruses. Proc Natl Acad Sci USA. 106:22205-10. * corresponding authors
Ijiri, H., Coulibaly F., Nishimura G., Nakai D., Chiu E., Takenaka C., Ikeda K., Nakazawa H., Hamada N., Kotani E., Metcalf P., Kawamata S., and Mori H. (2009). Structure-based targeting of bioactive proteins into cypovirus polyhedra and application to immobilized cytokines for mammalian cell culture. Biomaterials. 30:4297-4308
Kang H., Coulibaly F., Clow F., Proft T. and Baker E.N. (2007). Isopeptide Bonds Stabilize Gram-positive Bacterial Pilus Structure and Assembly. Science 318:1625-8.
Letzel T.§, Coulibaly F.§, Rey F.A., Delmas B., Jagt E., van Loon A.A. and Mundt E. (2007). Molecular and Structural Bases for the Antigenicity of VP2 of Infectious Bursal Disease Virus. J. Virol. 81:12827-35 §first authors
Hyun J.K. §, Coulibaly F. §, Turner A.P., Baker E.N., Mercer A.A. and Mitra A.K. (2007). The Structure of a Putative Scaffolding Protein of Immature Poxvirus Particles by Electron Microscopy Suggests Similarity with Capsid Proteins of Large Icosahedral DNA Viruses. J. Virol. 81:11075-83. §first authors
Coulibaly F., Chiu E., Ikeda K., Gutmann S., Haebel P.W., Schulze-Briese C., Mori H. and Metcalf, P. (2007). The Molecular Organization of Cypovirus Polyhedra. Nature 446:97-101.
Coulibaly, F., Chevalier, C., Galloux, M., Da Costa, B., Lepault, J., Delmas, B. and Rey, F.A. (2006). Les relations structurales entre birnavirus et autres virus icosaédriques à ARN. Virologie 10(3):233-235.
Coulibaly F., Chevalier C., Gutsche I., Pous J., Navaza J., Bressanelli S., Delmas B. and Rey F.A. (2005). The Birnavirus Crystal Structure Reveals Structural Relationships among Icosahedral Viruses. Cell 120:761-772.