John Briggs and Hans-Georg Kräusslich
MMPU Group 2011 – 2018
Project Summary
HIV-1, the causative agent of AIDS, is a major human pathogen with >30 million infected people world-wide and several million deaths annually. More than twenty drugs targeting different stages of HIV replication are in clinical use, but there is still an urgent need for new antivirals. Rapid and wide-spread resistance and side effects of available drugs limit the currently available therapeutic arsenal. We apply a combination of virology with state-of-the-art electron microscopy methods to decipher the architecture and dynamic alterations of HIV during assembly and maturation. Applying these methods, we aim to (i) understand how the virus develops resistance against existing drugs, (ii) how novel drugs and drug candidates work and (iii) obtain sufficiently detailed structural information to inform structure-based drug design of compounds blocking formation of the virion.
Background
Assembly and budding of HIV is directed by the viral Gag polyprotein. The virus is initially released as immature non-infectious particle consisting of mostly uncleaved Gag polyproteins. Extracellular maturation occurs after proteolytic processing of Gag at five different cleavage sites which is mediated by the virus-encoded, virion-associated protease (PR), translated as part of the Gag-Pol polyprotein. The immature virion consists of 2 – 3,000 Gag polyproteins arranged as spherical protein shell. Maturation leads to disassembly of the immature Gag layer, followed by a second assembly stage forming the mature cone-shaped capsid. Maturation and thus infectivity can be blocked by PR inhibitors, which are a mainstay of current anti-HIV therapy. Assembly of the immature as well as of the mature structure appear to be good antiviral targets, though only few compounds have been reported so far to affect these stages. The virion protein shell constitutes an irregular, heterogeneous structure, which makes structural analysis challenging. Cryo electron microscopy in combination with virology techniques is particularly suitable to overcome structural heterogeneity and to provide a higher resolution picture of the structures of the immature and mature virions as well as their intermediates. By doing so, these approaches will also provide the basis for an understanding of resistance to current drugs, define the mechanism of new compounds and identify new targets for structure-based drug development.
Research Goals
Research Goal 1: PI resistance
Protease inhibitors (PI) have been a key factor in the success of HIV therapy, but as with all clinically used drugs resistance has been observed. Compared to reverse transcriptase inhibitors, resistance against PI is mechanistically less well understood, but is believed to be a stepwise process involving resistance mutations in PR and compensatory mutations in PR and its cleavage sites as well as at other positions in Gag.
It is the aim of this collaborative work to determine the underlying molecular and structural mechanism and to decipher the complex interplay of substrate and enzyme mutations in PI resistance. Combined expertise in virology, biochemistry and structural biology is essential to uncover the complex mechanisms of PI resistance development and the role of substrate-enzyme co-evolution.
Research Goal 2: Assembly inhibitors
Assembly of both the immature and the mature lattice appear to be good targets for antiviral agents, as multiple weak interactions must cooperate to achieve the ordered protein shell. Disruption of only some interactions may be sufficient to affect infectivity since both stabilization and destabilization of the lattice seem to reduce specific infectivity of the virus.
Candidate compounds affecting assembly of either the immature or the mature protein shell (or both) have been developed by us and others, and some of these compounds have been shown to be active in tissue culture. However, detailed structural and functional analysis of their inhibitory mechanism and consequent improvement of their activity is now required to further develop such compounds into the clinic.
We are carrying out this analysis using virology, biochemistry and structural biology methods to explore the effect of inhibitors on virus assembly and on in vitro systems mimicking virus assembly.
Research Goal 3: Structure to inform drug design
To understand the assembly of the immature virus and to development new compounds directed against this structure, it is important to obtain a high-resolution structural view of the immature virus. The heterogeneous nature of the immature virus structure makes it difficult to study by conventional structural biology methods, and no high-resolution structure is currently available. We aim to resolve the structure of the immature virus to high resolution. To achieve this we are studying immature virus particles using cryo-electron tomography and sub-tomogram averaging methods.
In parallel we are developing protein constructs which can be assembled in vitro into structures which mimic the arrangement in the immature virus. Such in vitro assembled structures are potentially more regular and therefore more appropriate for high-resolution structural study. The structural data will represent a basis for the development of new molecules to inhibit assembly of the immature lattice (assembly inhibitors), and to inhibit PR mediated disassembly of the lattice (maturation inhibitors).
Selected publications
Maturation of the matrix and viral membrane of HIV-1.
Qu K, Ke Z, Zila V, Anders-Össwein M, Glass B, Mücksch F, Müller R, Schultz C, Müller B, Kräusslich HG, Briggs JAG. Science. 2021 Aug 6;373(6555):700-704. doi: 10.1126/science.abe6821. PMID: 34353956
Structure and architecture of immature and mature murine leukemia virus capsids.
Qu K, Glass B, Doležal M, Schur FKM, Murciano B, Rein A, Rumlová M, Ruml T, Kräusslich HG, Briggs JAG.
Proc Natl Acad Sci U S A. 2018 Dec 11;115(50):E11751-E11760. doi: 10.1073/pnas.1811580115. Epub 2018 Nov 26. PMID:30478053
High-resolution structures of HIV-1 Gag cleavage mutants determine structural switch for virus maturation.
Mattei S, Tan A, Glass B, Müller B, Kräusslich HG, Briggs JAG.
Proc Natl Acad Sci U S A. 2018 Oct 2;115(40):E9401-E9410. doi: 10.1073/pnas.1811237115. Epub 2018 Sep 14. PMID:30217893
The structure and flexibility of conical HIV-1 capsids determined within intact virions.
Mattei S, Glass B, Hagen WJ, Kräusslich HG, Briggs JA.
Science. 2016 Dec 16;354(6318):1434-1437.
PMID: 27980210
An atomic model of HIV-1 capsid-SP1 reveals structures regulating assembly and maturation.
Schur FK, Obr M, Hagen WJ, Wan W, Jakobi AJ, Kirkpatrick JM, Sachse C, Kräusslich HG, Briggs JA.
Science. 2016 Jul 29;353(6298):506-8. doi: 10.1126/science.aaf9620. Epub 2016 Jul 14.
PMID: 27417497
New hardware and workflows for semi-automated correlative cryo-fluorescence and cryo-electron microscopy/tomography.
Schorb M, Gaechter L, Avinoam O, Sieckmann F, Clarke M, Bebeacua C, Bykov YS, Sonnen AF, Lihl R, Briggs JA.
J Struct Biol. 2016 Jun 28. pii: S1047-8477(16)30135-6. doi: 10.1016/j.jsb.2016.06.020. [Epub ahead of print] PMID: 27368127
Nucleic Acid Binding by Mason-Pfizer Monkey Virus CA Promotes Virus Assembly and Genome Packaging.
Füzik T, Píchalová R, Schur FK, Strohalmová K, Křížová I, Hadravová R, Rumlová M, Briggs JA, Ulbrich P, Ruml T.
J Virol. 2016 Apr 14;90(9):4593-603. doi: 10.1128/JVI.03197-15. Print 2016 May 1. PMID: 26912613
A saposin-lipoprotein nanoparticle system for membrane proteins.
Frauenfeld J, Löving R, Armache JP, Sonnen AF, Guettou F, Moberg P, Zhu L, Jegerschöld C, Flayhan A, Briggs JA, Garoff H, Löw C, Cheng Y, Nordlund P.
Nat Methods. 2016 Apr;13(4):345-51. doi: 10.1038/nmeth.3801. Epub 2016 Mar 7. PMID: 26950744
Retrovirus maturation-an extraordinary structural transformation.
Mattei S, Schur FK, Briggs JA.
Curr Opin Virol. 2016 Mar 21;18:27-35. doi: 10.1016/j.coviro.2016.02.008. [Epub ahead of print] Review. PMID: 27010119
The Structure of Immature Virus-Like Rous Sarcoma Virus Gag Particles Reveals a Structural Role for the p10 Domain in Assembly.
Schur FK, Dick RA, Hagen WJ, Vogt VM, Briggs JA.
J Virol. 2015 Oct;89(20):10294-302. doi: 10.1128/JVI.01502-15. Epub 2015 Jul 29. PMID: 26223638
Structural Analysis of the Roles of Influenza A Virus Membrane-Associated Proteins in Assembly and Morphology.
Chlanda P, Schraidt O, Kummer S, Riches J, Oberwinkler H, Prinz S, Kräusslich HG, Briggs JA.
J Virol. 2015 Sep 1;89(17):8957-66. doi: 10.1128/JVI.00592-15. Epub 2015 Jun 17. PMID:26085153
RNA and nucleocapsid are dispensable for mature HIV-1 capsid core assembly.
Mattei S, Flemming A, Anders-Össwein M, Kräusslich HG, Briggs JA, Müller B.
J Virol. 2015 Jul 15. pii: JVI.00750-15. [Epub ahead of print] PMID:26178992
Retroviral proteases and their roles in virion maturation.
Konvalinka J, Kräusslich HG, Müller B.
Virology. 2015 May;479-480:403-17. doi: 10.1016/j.virol.2015.03.021. Epub 2015 Mar 26. Review. PMID: 25816761
Triggering HIV polyprotein processing by light using rapid photodegradation of a tight-binding protease inhibitor.
Schimer J, Pávová M, Anders M, Pachl P, Šácha P, Cígler P, Weber J, Majer P, Řezáčová P, Kräusslich HG, Müller B, Konvalinka J.
Nat Commun. 2015 Mar 9;6:6461. doi: 10.1038/ncomms7461. PMID:25751579
Structure of the immature HIV-1 capsid in intact virus particles at 8.8 Å resolution.
Schur FK, Hagen WJ, Rumlová M, Ruml T, Müller B, Kräusslich HG, Briggs JA.
Nature. 2015 Jan 22;517(7535):505-8. doi: 10.1038/nature13838. Epub 2014 Nov 2.
PMID: 25363765
Induced maturation of human immunodeficiency virus.
Mattei S, Anders M, Konvalinka J, Kräusslich HG, Briggs JA, Müller B.
J Virol. 2014 Dec 1;88(23):13722-31. doi: 10.1128/JVI.02271-14. Epub 2014 Sep 17.
Cryo-electron microscopy of tubular arrays of HIV-1 Gag resolves structures essential for immature virus assembly.
Bharat, T.A., Castillo Menendez, L.R., Hagen, W.J., Lux, V., Igonet, S., Schorb, M., Schur, F.K., Krausslich, H.G. & Briggs, J.A.
Proc Natl Acad Sci U S A. 2014 May 19. pii: 201401455. Europe PMC
The NC domain of Gag is dispensable for actin incorporation into HIV-1 and for association of viral budding sites with cortical F-actin.
Stauffer S1, Rahman SA, de Marco A, Carlson LA, Glass B, Oberwinkler H, Herold N, Briggs JA, Müller B, Grünewald K, Kräusslich HG._
J Virol. 2014 Apr 30. [Epub ahead of print]_
Role of the SP2 Domain and Its Proteolytic Cleavage in HIV-1 Structural Maturation and Infectivity
de Marco, A., Heuser, A.M., Glass, B., Krausslich, H.G., Muller, B. & Briggs, J.A.
J Virol. 2012 Dec;86(24):13708-16. doi: 10.1128/JVI.01704-12. Epub 2012 Oct 10
The molecular architecture of HIV.
Briggs, J. A. Krausslich, H. G.
J Mol Biol 2011 410, 491-500. (Review)
Structural analysis of HIV-1 maturation using cryo-electron tomography.
de Marco, A., Muller, B., Glass, B., Riches, J. D., Krausslich, H. G. Briggs, J. A.
PLoS Pathog 2010 6, e1001215.
Cryo electron tomography of native HIV-1 budding sites.
Carlson, L. A., de Marco, A., Oberwinkler, H., Habermann, A., Briggs, J. A., Krausslich, H. G. Grunewald, K.
PLoS Pathog 2010 6, e1001173.
Structure and assembly of immature HIV.
Briggs, J. A., Riches, J. D., Glass, B., Bartonova, V., Zanetti, G. Krausslich, H. G.
Proc Natl Acad Sci U S A 2009 106, 11090-5.
Group Members
in alphabetical order
Tanmay Bharat, PhD, Postdoc
John Briggs, PhD
MMPU Group Leader
now: Director at the Max-Planck-Institut for Biochemistry, Munich, Germany.
Annica Flemming, PhD Student
Hans-Georg Kräusslich, MD
MMPU Group Leader
Director of the Department of Infectious Diseases, Virology, Heidelberg University Medical Center
Alex de Marco, PhD Student (EMBL International PhD Programme – EIPP)
Simone Mattei, PhD Student (EMBL International PhD Programme – EIPP)
Luis Castillo Menendez, PhD Student
Barbara Müller, PhD, Group Leader
Martin Obr, PhD, Postdoc
Kun Qu, Postdoc
Rene Sahm, PhD Student
Florian Schur, PhD Student (EMBL International PhD Programme – EIPP)
Andreas Sonnen, MD, Postdoc
Aaron Tan, PhD Student (EMBL International PhD Programme – EIPP)
