Generation of a Family-specific Phage Library of Llama Single Chain Antibody Fragments That Neutralize HIV-1

Abstract

Recently, we described llama antibody fragments (VHH) that can neutralize human immunodeficiency virus, type 1 (HIV-1). These VHH were obtained after selective elution of phages carrying an immune library raised against gp120 of HIV-1 subtype B/C CN54 with soluble CD4. We describe here a new, family-specific approach to obtain the largest possible diversity of related VHH that compete with soluble CD4 for binding to the HIV-1 envelope glycoprotein. The creation of this family-specific library of homologous VHH has enabled us to isolate phages carrying similar nucleotide sequences as the parental VHH. These VHH displayed varying binding affinities and neutralization phenotypes to a panel of different strains and subtypes of HIV-1. Sequence analysis of the homologs showed that the C-terminal three amino acids of the CDR3 loop were crucial in determining the specificity of these VHH for different subtype C HIV-1 strains. There was a positive correlation between affinity of VHH binding to gp120 of HIV-1 IIIB and the breadth of neutralization of diverse HIV-1 envelopes. The family-specific approach has therefore allowed us to better understand the interaction of the CD4-binding site antibodies with virus strain specificity and has potential use for the bioengineering of antibodies and HIV-1 vaccine development.

HIV sexual transmission and microbicides

“…Other advantages of VHHs include (1) preference for cleft recognition and for binding into active sites [200,201], (2) ease of cloning and expression in bacteria and yeast [202] and (3) excellent stability and solubility [203,204]. These characteristics have led to the development of camelid nanobodies for a number of biological targets [205][206][207][208][209][210][211][212][213], including HIV [199,214,215].…”

Straightforward Selection of Broadly Neutralizing Single-Domain Antibodies Targeting the Conserved CD4 and Coreceptor Binding Sites of HIV-1 gp120

“…Thus, because of their small size and their tendency to bind cavities, they have been proposed as good neutralizing antibody candidates. If CD4BS sdAbs with good neutralizing capacities have already been isolated (13,14), to our knowledge, no CoRBS sdAb has yet been discovered to date.…”

HIV vaccines: current status worldwide and in Africa

“…Plasma samples were screened against a panel of viruses, and the donors of broadly neutralizing antibodies (bnAbs) were selected for harvest of PBMC and attempted cloning of the antibodies [16]. Additional antibodies with similar breadth and potency are now being identified by other groups using different methods [12,13,17].…”