TY - JOUR
T1 - Profound structural conservation of chemically cross-linked HIV-1 envelope glycoprotein experimental vaccine antigens
AU - Martin, Gregory M.
AU - Russell, Rebecca A.
AU - Mundsperger, Philip
AU - Harris, Scarlett
AU - Jovanoska, Lu
AU - Trajano, Luiza Farache
AU - Schiffner, Torben
AU - Fabian, Katalin
AU - Tolazzi, Monica
AU - Scarlatti, Gabriella
AU - McFarlane, Leon
AU - Cheeseman, Hannah
AU - Aldon, Yoann
AU - Schermer, Edith E.
AU - Breemen, Marielle
AU - Sliepen, Kwinten
AU - Katinger, Dietmar
AU - Kunert, Renate
AU - Sanders, Rogier W.
AU - Shattock, Robin
AU - Ward, Andrew B.
AU - Sattentau, Quentin J.
N1 - Funding Information: We thank John Mascola, Peter Kwong, Dennis Burton, Michel Nussenzweig, Mark Connors, and James Robinson for donating antibodies and other reagents either directly or through the NIH AIDS Reagents Program. We thank Dennis Burton and the IAVI Neutralizing Antibody Consortium for reagents. Q.J.S. is a Jenner Institute Investigator and James Martin School Senior Fellow. This research was supported by The European Union H2020 European AIDS Vaccine Initiative (EAVI2020 https://ec.europa.eu/programmes/horizon2020/ ) award No. 681137 (R.R., P.M., K.F., M.T., G.S., H.C., Y.A., M.B., R.W.S., R.S., Q.J.S.), Fondation Dormeur, Vaduz (G.S. and Q.J.S.), NIH grant UM1AI100663 (A.B.W.), UM1AI144462 (A.B.W.), the Bill and Melinda Gates Foundation grants OPP1115782 (A.B.W.), OPP1170236 (A.B.W.). Funding Information: We thank John Mascola, Peter Kwong, Dennis Burton, Michel Nussenzweig, Mark Connors, and James Robinson for donating antibodies and other reagents either directly or through the NIH AIDS Reagents Program. We thank Dennis Burton and the IAVI Neutralizing Antibody Consortium for reagents. Q.J.S. is a Jenner Institute Investigator and James Martin School Senior Fellow. This research was supported by The European Union H2020 European AIDS Vaccine Initiative (EAVI2020 https://ec.europa.eu/programmes/horizon2020/ ) award No. 681137 (R.R., P.M., K.F., M.T., G.S., H.C., Y.A., M.B., R.W.S., R.S., Q.J.S.), Fondation Dormeur, Vaduz (G.S. and Q.J.S.), NIH grant UM1AI100663 (A.B.W.), UM1AI144462 (A.B.W.), the Bill and Melinda Gates Foundation grants OPP1115782 (A.B.W.), OPP1170236 (A.B.W.). Publisher Copyright: © 2023, The Author(s).
PY - 2023/12/1
Y1 - 2023/12/1
N2 - Chemical cross-linking is used to stabilize protein structures with additional benefits of pathogen and toxin inactivation for vaccine use, but its use has been restricted by the potential for local or global structural distortion. This is of particular importance when the protein in question requires a high degree of structural conservation for inducing a biological outcome such as the elicitation of antibodies to conformationally sensitive epitopes. The HIV-1 envelope glycoprotein (Env) trimer is metastable and shifts between different conformational states, complicating its use as a vaccine antigen. Here we have used the hetero-bifunctional zero-length reagent 1-Ethyl-3-(3-Dimethylaminopropyl)-Carbodiimide (EDC) to cross-link two soluble Env trimers, selected well-folded trimer species using antibody affinity, and transferred this process to good manufacturing practice (GMP) for experimental medicine use. Cross-linking enhanced trimer stability to biophysical and enzyme attack. Cryo-EM analysis revealed that cross-linking retained the overall structure with root-mean-square deviations (RMSDs) between unmodified and cross-linked Env trimers of 0.4–0.5 Å. Despite this negligible distortion of global trimer structure, we identified individual inter-subunit, intra-subunit, and intra-protomer cross-links. Antigenicity and immunogenicity of the trimers were selectively modified by cross-linking, with cross-linked ConS retaining bnAb binding more consistently than ConM. Thus, the EDC cross-linking process improves trimer stability whilst maintaining protein folding, and is readily transferred to GMP, consistent with the more general use of this approach in protein-based vaccine design.
AB - Chemical cross-linking is used to stabilize protein structures with additional benefits of pathogen and toxin inactivation for vaccine use, but its use has been restricted by the potential for local or global structural distortion. This is of particular importance when the protein in question requires a high degree of structural conservation for inducing a biological outcome such as the elicitation of antibodies to conformationally sensitive epitopes. The HIV-1 envelope glycoprotein (Env) trimer is metastable and shifts between different conformational states, complicating its use as a vaccine antigen. Here we have used the hetero-bifunctional zero-length reagent 1-Ethyl-3-(3-Dimethylaminopropyl)-Carbodiimide (EDC) to cross-link two soluble Env trimers, selected well-folded trimer species using antibody affinity, and transferred this process to good manufacturing practice (GMP) for experimental medicine use. Cross-linking enhanced trimer stability to biophysical and enzyme attack. Cryo-EM analysis revealed that cross-linking retained the overall structure with root-mean-square deviations (RMSDs) between unmodified and cross-linked Env trimers of 0.4–0.5 Å. Despite this negligible distortion of global trimer structure, we identified individual inter-subunit, intra-subunit, and intra-protomer cross-links. Antigenicity and immunogenicity of the trimers were selectively modified by cross-linking, with cross-linked ConS retaining bnAb binding more consistently than ConM. Thus, the EDC cross-linking process improves trimer stability whilst maintaining protein folding, and is readily transferred to GMP, consistent with the more general use of this approach in protein-based vaccine design.
UR - http://www.scopus.com/inward/record.url?scp=85165244101&partnerID=8YFLogxK
U2 - https://doi.org/10.1038/s41541-023-00696-w
DO - https://doi.org/10.1038/s41541-023-00696-w
M3 - Article
C2 - 37443366
SN - 1476-0584
VL - 8
JO - npj Vaccines
JF - npj Vaccines
IS - 1
M1 - 101
ER -