Program in Mathematical, Computational and Systems Biology, University of California, Irvine, California 92697, United States.
Department of Biomedical Engineering, University of California, Irvine, California 92697, United States.
ACS Synth Biol. 2024 Aug 16;13(8):2629-2634. doi: 10.1021/acssynbio.4c00370. Epub 2024 Jul 25.
We recently developed "autonomous hypermutation yeast surface display" (AHEAD), a technology that enables the rapid generation of potent and specific antibodies in yeast. AHEAD pairs yeast surface display with an error-prone orthogonal DNA replication system (OrthoRep) to continuously and rapidly mutate surface-displayed antibodies, thereby enabling enrichment for stronger binding variants through repeated rounds of cell growth and fluorescence-activated cell sorting. AHEAD currently utilizes a standard galactose induction system to drive the selective display of antibodies on the yeast surface. However, achieving maximal display levels can require up to 48 h of induction. Here we report an updated version of the AHEAD platform that utilizes a synthetic β-estradiol-induced gene expression system to regulate the surface display of antibodies and find that induction is notably faster in achieving surface display for both our AHEAD system and traditional yeast surface display from nuclear plasmids that do not hypermutate. The updated AHEAD platform was fully functional in repeated rounds of evolution to drive the rapid evolution of antibodies.
我们最近开发了“自主超突变酵母表面展示”（AHEAD）技术，该技术可在酵母中快速产生高效且特异性的抗体。AHEAD 将酵母表面展示与易错的正交 DNA 复制系统（OrthoRep）相结合，持续快速地突变表面展示的抗体，从而通过反复的细胞生长和荧光激活细胞分选来富集更强结合的变体。AHEAD 当前利用标准半乳糖诱导系统来驱动抗体在酵母表面的选择性展示。然而，要实现最大的展示水平，可能需要长达 48 小时的诱导。在这里，我们报告了 AHEAD 平台的更新版本，该版本利用合成的β-雌二醇诱导基因表达系统来调节抗体的表面展示，并且发现诱导在实现我们的 AHEAD 系统和传统的核质体酵母表面展示的表面展示方面都明显更快，而后者不会发生超突变。更新后的 AHEAD 平台在反复的进化循环中完全有效，可快速进化抗体。
