Anti-HIV type 1 Glycoprotein 120 Polyclonal antibody

As a step toward the development of variant forms of Env with enhanced immunogenic properties, we have expressed the glycoprotein in the yeast surface display system in a form that can be subjected to random mutagenesis followed by screening for forms with enhanced binding to germline antibodies. To optimize the expression and immunogenicity of the yeast-displayed Env protein, we tested different approaches for cell wall anchoring, expression of gp120 and gp140 Env from different viral strains, the effects of introducing mutations designed to stabilize Env, and the effects of procedures for altering N-linked glycosylation of Env. We find that diverse forms of HIV envelope glycoprotein can be efficiently expressed at the yeast cell surface and that gp140 forms of Env are effectively cleaved by Kex2p, the yeast furin protease homolog. Multiple yeast-displayed gp120 and gp140 proteins are capable of binding to antibodies directed against the V3-variable loop, CD4 binding site, and gp41 membrane-proximal regions, including some antibodies whose binding is known to depend on Env conformation and N-linked glycan. Based on antibody recognition and sensitivity to glycosidases, yeast glycosylation patterns partially mimic high mannose-type N-glycosylation in mammalian cells. However, yeast-displayed Env is not recognized by some anti-Env antibodies sensitive to quaternary structure, suggesting either that the displayed protein exists in a monomeric state or that for these antibodies, yeast glycosylation in certain regions hinders recognition or access. Consistent with studies in other systems, reconstructed predicted unmutated precursors to anti-Env antibodies exhibit little affinity for the yeast-displayed envelope protein.

Inducing a long-lasting as well as broad and potent immune response by generating broadly neutralizing antibodies is a major goal and at the same time the main challenge of preventive HIV-1 vaccine design. Immunization with soluble, stabilized and native-like envelope (Env) glycoprotein so far only led to low neu-tralization breadth and displayed low immunogenicity. A promising approach to generate a potent immune response is the presentation of Env on the surface of nanoparticles. In this review, we will focus on two key processes essential for the induction of immune response that can be addressed by specific features of nanoparticulate carriers: first, the trafficking to and within distinct compartments of the lymph node, and second, the use of multivalent Env display allowing for high avidity interactions. To optimize these pivotal steps critical design criteria should be considered for the presentation of Env on nanoparticles. These include an optimal particle size below 100 nm, distances between two adjacent Env antigens of approximately 10-15 nm, an appropriate orientation of Env, and finally, the stability of both the Env attachment and the nanoparticle platform. Hence, an interdisciplinary approach that combines a suitable delivery system and a straightforward presentation of the Env antigen may have the potential to drive the immune response towards increased breadth and potency.