Heavy-chain-only antibodies (HCAbs), which are devoid of light chains, have been found
naturally occurring in various species including camelids and cartilaginous fish. The
variable regions of the HCAbs (VHHs) are known as the minimal natural antigen recognition
units. Most immunogenetic and functional studies of HCAbs are based on case studies or a
limited number of low-throughput sequencing data. A complete picture derived from more
abundant high-throughput sequencing (HTS) data can help us gain deeper insights. In this
study, we cloned and sequenced the full-length coding region of VHHs from non-immunized
Alpaca (Vicugna pacos) via HTS. A new pipeline was developed to conduct an in-depth analysis
of the HCAb repertoires. Various critical features, including the length distribution of
complementarity-determining region 3 (CDR3), V(D)J usage, VJ pairing, germline-specific
mutation rate, and germline-specific scoring profiles (GSSPs), were systematically
characterized. The quantitative data show that V(D)J usage and VHH recombination are highly
biased. Interestingly, we found that the average CDR3 length of classical VHHs is longer
than that of non-classical ones, whereas the mutation rates are similar in both kinds of
VHHs. Finally, GSSPs were built to quantitatively describe and compare sequences that
originate from each VJ pair. Overall, this study presents a comprehensive landscape of the
HCAb repertoire, which can provide useful guidance for the modeling of somatic hypermutation
and the design of novel functional VHHs or VHH repertoires via evolutionary profiles.
Zhui Tu, Xiaoqiang Huang, Jinheng Fu, Na Hu, Wei Zheng, Yanping Li, Yang Zhang.
Landscape of Variable Domain of Heavy-chain-only Antibody Repertoire from Alpaca.
Immunology, 161: 53-65 (2020).