Lu Yonggang, Ikawa Masahito, Tang Shaogeng
Premium Research Institute for Human Metaverse Medicine, Osaka University, Suita, Osaka 565-0871, Japan.
Research Institute for Microbial Diseases, Osaka University, Suita, Osaka 565-0871, Japan.
Proc Natl Acad Sci U S A. 2025 Mar 11;122(10):e2425952122. doi: 10.1073/pnas.2425952122. Epub 2025 Mar 5.
Sperm IZUMO1 binds to egg JUNO, and this interaction is essential for mammalian fertilization. Isolated from a female mouse immunized with syngeneic sperm, the antisperm antibody OBF13 recognizes IZUMO1 and inhibits murine fertilization. How OBF13 interferes with sperm-egg interactions was unknown. Here, we present the X-ray crystal structure of IZUMO1 in complex with OBF13. OBF13 binds to the apex of the four-helix domain of IZUMO1, distant from the JUNO-binding site. Our crystal structure of OBF13-bound IZUMO1 resembles apo-IZUMO1 and differs from the structure of IZUMO1 in complex with JUNO. We identify that OBF13 carries a low level of somatic hypermutation, and through deep mutational scanning, we engineer an affinity-enhanced OBF13 variant. This OBF13 variant single-chain fragment variable decreases the apparent affinity of IZUMO1 for membrane-bound murine JUNO and blocks the binding of acrosome-reacted sperm to eggs, thereby preventing fertilization. We propose allostery between the OBF13 epitope and the JUNO-binding site. OBF13 inhibits a conformational change in IZUMO1, preventing fusion-competent sperm from adhering to murine eggs during fertilization. Surprisingly, murine IZUMO1 binds to hamster JUNO with an affinity ~20-fold higher than to murine JUNO. The decreased affinity caused by OBF13 of murine IZUMO1 for hamster JUNO is sufficient for murine sperm to bind to and fuse with hamster eggs. Our studies provide a structural and mechanistic framework for species-specific, allosteric inhibition of IZUMO1 by a naturally occurring antisperm antibody and offer insights into the development of immunocontraceptives.
精子IZUMO1与卵子JUNO结合，这种相互作用对哺乳动物受精至关重要。从用同种精子免疫的雌性小鼠中分离出的抗精子抗体OBF13可识别IZUMO1并抑制小鼠受精。OBF13如何干扰精卵相互作用尚不清楚。在此，我们展示了与OBF13结合的IZUMO1的X射线晶体结构。OBF13结合到IZUMO1四螺旋结构域的顶端，远离JUNO结合位点。我们得到的与OBF13结合的IZUMO1的晶体结构类似于无配体的IZUMO1，与和JUNO结合的IZUMO1结构不同。我们发现OBF13存在低水平的体细胞超突变，并通过深度突变扫描构建了一个亲和力增强的OBF13变体。这种OBF13变体单链可变片段降低了IZUMO1对膜结合型小鼠JUNO的表观亲和力，并阻断顶体反应精子与卵子的结合，从而阻止受精。我们提出OBF13表位与JUNO结合位点之间存在变构作用。OBF13抑制IZUMO1的构象变化，防止具有融合能力的精子在受精过程中黏附到小鼠卵子上。令人惊讶的是，小鼠IZUMO1与仓鼠JUNO的结合亲和力比对小鼠JUNO的亲和力高约20倍。OBF13导致的小鼠IZUMO1对仓鼠JUNO亲和力的降低足以使小鼠精子与仓鼠卵子结合并融合。我们的研究为天然抗精子抗体对IZUMO1进行物种特异性变构抑制提供了结构和机制框架，并为免疫避孕的发展提供了见解。
