Transient opening of trimeric prefusion RSV F proteins
Gilman MSA, Furmanova-Hollenstein P, Pascual G, van ‘t Wout AB, Langedijk JPM, McLellan JS. Nat Commun. 2015 May 8;10(1):2105. Full text PDF
The respiratory syncytial virus F glycoprotein is a class I fusion protein that mediates viral entry and is the target of a number of clinical interventions currently in development for RSV. This manuscript demonstrates for the first time that the prefusion RSV F trimer transiently dissociates. The serendipitous discovery of a monoclonal antibody that favors trimer dissociation, CR9501, allowed us to determine that the prefusion RSV F trimer is in a monomer–trimer equilibrium in solution and when expressed on the surface of mammalian cells. Structural studies reveal that the prefusion F trimer samples at least two conformations, one of which reduced contacts between prefusion F monomers and might favor trimer opening. These results also have implications for RSV vaccine design, since several vaccines in clinical development are based on trimeric prefusion-stabilized F antigens.
Iterative screen optimization maximizes the efficiency of macromolecular crystallization
Jones HJ, Wrapp D, Gilman MSA, Battles MB, Wang N, Sacerdote S, Chuang G-Y, Kwong PD, McLellan JS. Acta Cryst. (2019). F75, 123-131. PMID: 30713164
The rate-limiting step in most crystallographic experiments is obtaining diffraction-quality macromolecular crystals. In this manuscript we describe the technique of Iterative Screen Optimization (ISO), which seeks to maximize the efficiency of crystallization. To take full advantage of this method we designed a novel 96-well crystallization screen, composed of common, inexpensive chemical reagents. This novel screen is used to prepare conventional high-throughput crystallization plates. After several days of incubation, each drop in the plates is assigned a user-generated score based on its qualitative appearance. Based on these scores, the precipitant concentrations making up each drop are optimized and the entire high-throughput crystallization screen is automatically reformulated and re-dispensed. When this process is performed iteratively, these screens gradually become tailored toward their given crystallization target, maximizing the efficiency of the chemical reagents that make up the screen. We went on to demonstrate the utility of this novel method by using it to rapidly crystallize a panel of six diverse proteins. It is our hope that this technique will become widely adopted by crystallographers working with recalcitrant macromolecules.
Structural basis for recognition of the central conserved region of RSV G by neutralizing human antibodies
Jones HG, Ritschel T, Pascual G, Brakenhoff JPJ, Keogh E, Furmanova-Hollenstein P, Lanckacker E, Wadia JS, Gilman MSA, Williamson RA, Roymans D, van 't Wout AB, Langedijk JP, McLellan JS. PLoS Pathog. 2018 Mar 6;14(3):e1006935. PMID: 29509814
The attachment glycoprotein (RSV G) purportedly binds to CX3CR1 on human airway epithelial cells to mediate viral attachment and subsequent infection. This manuscript describes the characterization of two human antibodies that bind to the central conserved region of RSV G, potently neutralize both subtypes of RSV, and protect RSV-challenged animals from severe disease. We also determined high-resolution crystal structures of each antibody in complex with the central conserved region of RSV G to gain a better understanding of how these antibodies bind to RSV G and how they neutralize the virus. Comparison of these structures with the structure of fractalkine (CX3CL1 – the natural ligand of CX3CR1) suggests that RSV G likely binds to CX3CR1 in a mode that is distinct from that of fractalkine. Collectively, these results build on recent studies demonstrating the importance of RSV G in antibody-mediated protection, and the structural information should guide the development of new vaccines as well as the investigation of the structural interactions between RSV G and CX3CR1 that mediate viral attachment.
Infants infected with respiratory syncytial virus generate potent neutralizing antibodies that lack somatic hypermutation
Goodwin E, Gilman MSA, Wrapp D, Chen M, Ngwuta JO, Moin SM, Bai P, Sivasubramanian A, Connor RI, Wright PF, Graham BS, McLellan JS, Walker LM. Immunity 2018; 48(2):339-349.e5.
Respiratory syncytial virus (RSV) is a leading cause of infant mortality, and there are currently no licensed vaccines to protect this vulnerable population. This manuscript describes the isolation and characterization of more than 450 RSV fusion glycoprotein (F)-specific antibodies from 7 RSV-infected infants. These studies revealed that infant antibody repertoires are highly focused compared to those of adults, and that antibodies directed against two dominant sites utilize convergent sequence features for recognition of RSV F. A subset of antibodies targeting one of these sites displayed potent neutralizing activity despite lacking somatic mutations, suggesting that expansion of these B cells in infants may be possible with suitably designed vaccine antigens. Structural studies revealed the molecular determinants for the convergent recognition of both immunodominant sites, providing a framework for the rational design of age-specific RSV vaccines.
Battles MB, Más V, Olmedillas E, Cano O, Vázquez M, Rodríguez L, Melero JA, McLellan JS.
Structure and immunogenicity of prefusion-stabilized human metapneumovirus F glycoprotein.
Nat Commun. 2017 Nov 16;8(1):1528.
Tian D, Battles MB, Moin SM, Chen M, Modjarrad K, Kumar A, Kanekiyo M, Graepel KW, Taher NM, Hotard AL, Moore ML, Zhao M, Zheng ZZ, Xia NS, McLellan JS, Graham BS.
Structural basis of respiratory syncytial virus subtype-dependent neutralization by an antibody targeting the fusion glycoprotein.
Nat Commun. 2017 Nov 30;8(1):1877.
Pallesen J, Wang N, Corbett KS, Wrapp D, Kirchdoerfer RN, Turner HL, Cottrell CA, Becker MM, Wang L, Shi W, Kong WP, Andres EL, Kettenbach AN, Denison MR, Chappell JD, Graham BS, Ward AB, McLellan JS.
Immunogenicity and structures of a rationally designed prefusion MERS-CoV spike antigen
PNAS 2017 Aug 29;114(35):E7348-E7357.
Rossey I, Gilman MS, Kabeche SC, Sedeyn K, Wrapp D, Kanekiyo M, Chen M, Mas V, Spitaels J, Melero JA, Graham BS, Schepens B, McLellan JS, Saelens X.
Potent single-domain antibodies that arrest respiratory syncytial virus fusion protein in its prefusion state.
Nat Commun. 2017 Feb 13;8:14158.
Gilman MS, Castellanos CA, Chen M, Ngwuta JO, Goodwin E, Moin SM, Mas V, Melero JA, Wright PF, Graham BS, McLellan JS, Walker LM.
Rapid profiling of RSV antibody repertoires from the memory B cells of naturally infected adult donors.
Sci Immunol. 2016 Dec 16;1(6). pii: eaaj1879.
Más V, Rodriguez L, Olmedillas E, Cano O, Palomo C, Terron MC, Luque D, Melero JA, McLellan JS.
Engineering, structure and immunogenicity of the human metapneumovirus F protein in the postfusion conformation.
PLoS Pathog. 2016 Sep 9;12(9):e1005859.
Misasi J, Gilman MS, Kanekiyo M, Gui M, Cagigi A, Mulangu S, Corti D, Ledgerwood JE, Lanzavecchia A, Cunningham J, Muyembe-Tamfun JJ, Baxa U, Graham BS, Xiang Y, Sullivan NJ, McLellan JS.
Structural and molecular basis for Ebola virus neutralization by protective human antibodies.
Science. 2016 Mar 18;351(6279):1343-6.
Kirchdoerfer RN, Cottrell CA, Wang N, Pallesen J, Yassine HM, Turner HL, Corbett KS, Graham BS, McLellan JS, Ward AB.
Pre-fusion structure of a human coronavirus spike protein.
Nature. 2016 Mar 3;531(7592):118-21.
Battles MB, Langedijk JP, Furmanova-Hollenstein P, Chaiwatpongsakorn S, Costello HM, Kwanten L, Vranckx L, Vink P, Jaensch S, Jonckers TH, Koul A, Arnoult E, Peeples ME, Roymans D, McLellan JS.
Molecular mechanism of respiratory syncytial virus fusion inhibitors.
Nat Chem Biol. 2016 Feb;12(2):87-93.
Krarup A, Truan D, Furmanova-Hollenstein P, Bogaert L, Bouchier P, Bisschop IJ, Widjojoatmodjo MN, Zahn R, Schuitemaker H, McLellan JS, Langedijk JP.
A highly stable prefusion RSV F vaccine derived from structural analysis of the fusion mechanism.
Nat Commun. 2015 Sep 3;6:8143
Gilman MS, Moin SM, Mas V, Chen M, Patel NK, Kramer K, Zhu Q, Kabeche SC, Kumar A, Palomo C, Beaumont T, Baxa U, Ulbrandt ND, Melero JA, Graham BS, McLellan JS.
Characterization of a prefusion-specific antibody that recognizes a quaternary, cleavage-dependent epitope on the RSV fusion glycoprotein.
PLoS Pathog. 2015 Jul 10; 11(7):e1005035.