A Role for Small Antibody Fragments to Bind and Neutralize HIV
The surface of the Human Immunodeficiency Virus (HIV) is studded with numerous copies of the glycoprotein Env. Each Env spike is composed of three copies of the proteins gp41, which sits in the viral membrane, and gp120, which rests on top of each gp41 molecule. Env is essential for HIV-mediated infection because the binding of gp120 to the T cell surface receptor CD4 initiates a conformational change in Env exposing the fusion peptide, which inserts into the T cell membrane and helps fuse the T cell and virus together. This makes Env an attractive target for designing therapeutic inhibitory antibodies. However, the complexities of the HIV surface proteins and the tight association of the virus and T cell during infection have hampered the identification of full-length antibodies with effective HIV neutralizing activity.
More recently, researchers have developed antibody derivatives, which are single domains from full-length antibodies or engineered protein sequences, capable of recognizing regions of Env. These derivatives are advantageous because of their smaller size, improved stability, reduced cost, and ease of production compared with their full-length counterparts. To better understand how two of these derivatives interact with Env on the surface of intact HIV particles, Sriram Subramaniam, Ph.D., in CCR’s Laboratory of Cell Biology, and colleagues determined the three-dimensional structures of each derivative bound to Env.
The researchers first investigated the structure of viruses expressing Env alone or in complex with A12, the variable region of a heavy-chain only llama antibody. They rapidly froze the samples then imaged them using electron microscopy. Multiple images were taken at varying angles and combined to recreate the three-dimensional arrangement of the Env proteins bound to A12 or not. In the A12-bound structure, the gp120 proteins were further apart and more density was observed than in the unbound version. When the scientists overlaid detailed structures of A12-bound Env determined by X-ray crystallography, A12 occupied the area of excess density. Additionally, A12 binding caused an outward rotation of the bound gp120 proteins. This structure is similar to that seen for the complex of Env with the full-length neutralizing antibody b12, which the researchers determined in previous studies and described as a “partially open” conformation. The research team showed that A12 and b12, but not another antibody, VRC01, that interacts with the same region of gp120, required the rotation of gp120 to accommodate their binding.
The second antibody derivative the investigators chose to study was m36, the variable region of a human antibody heavy chain. m36, originally described by Dimiter Dimitrov in CCR’s Nanobiology Program, also binds to gp120 but at a region distinct from A12. The researchers demonstrated that m36 binding to gp120 was enhanced in the presence of soluble CD4. This type of binding mirrors that of the CD4-induced antibody 17b. Likewise, the scientists found that the three-dimensional structures of both 17b-bound Env and Env bound to m36 showed large movements of gp120, a structure they termed the “open” conformation of Env. Because a high-resolution structure of m36 is not yet available, the researchers determined the structure of Env bound to both m36 and soluble CD4. By subtracting out the structure of Env interacting with soluble CD4 alone, they determined that m36 binds to the base of one loop of gp120, a location similar to that bound by 17b.
Taken together, these results show that at least these two antibody derivatives, A12 and m36, can cause structural rearrangements in Env analogous to those of the full-length neutralizing antibodies, b12 and 17b, respectively. Thus, these derivatives may have potential therapeutic applications and deserve further study.Summary Posted: 02/2013
Meyerson JR, Tran EE, Kuybeda O, Chen W, Dimitrov DS, Gorlani A, Verrips T, Lifson JD, Subramaniam S. Molecular structures of trimeric HIV-1 Env in complex with small antibody derivatives.PNAS, 110(2):513-8, Jan. 8, 2013. PubMed Link