Harding Maxim D, Jackson Mark A, Gilding Edward K, Yap Kuok, Craik David J, Sainsbury Frank, Lawrence Nicole
Institute for Molecular Bioscience, Australian Research Council Centre of Excellence for Innovations in Peptide and Protein Science, The University of Queensland, Brisbane, Queensland, Australia.
Centre for Cell Factories and Biopolymers, Institute for Biomedicine and Glycomics, Griffith University, Nathan, Queensland, Australia.
Microb Biotechnol. 2025 Jun;18(6):e70158. doi: 10.1111/1751-7915.70158.
New therapeutics are urgently needed to curb the spread of drug-resistant diseases. Bioactive peptides (BAPs), including antimicrobial peptides, are emerging as an exciting new class of compounds with advantages over current drug modalities, especially small molecule drugs that are prone to resistance development. Here, we evaluated a bacteriophage P22 virus-like particle (VLP) system where BAPs are encapsulated as fusion proteins with the P22 scaffold protein (SP) within self-assembling protein cages in Escherichia coli. Representative peptides from three structurally distinct classes of BAPs were successfully encapsulated into P22 VLPs at high cargo to VLP coat protein (CP) ratios that corresponded to interactions between the compact electropositive structures of the SP-BAPs and electronegative regions on the inward facing surface of CP subunits. However, high loading densities did not correspond to improved SP-BAP yields. An unexpected finding of this study was that while encapsulation alleviated negative effects of SP-BAPs on E. coli growth, the P22 scaffold protein acted as a sufficient fusion partner for accumulating BAPs, and co-expression of the CP did not further improve SP-BAP yields. Nevertheless, encapsulation in VLPs provided a useful first step in the purification pipeline for producing both linear and cyclic recombinant (r)BAPs that were functionally equivalent to their synthetic counterparts. Further efforts to optimise expression ratios of CP to SP-BAP fusions will be required to realise the full potential of encapsulation for protecting expression hosts and maximising rBAP yields.
迫切需要新的治疗方法来遏制耐药性疾病的传播。生物活性肽（BAPs），包括抗菌肽，正成为一类令人兴奋的新型化合物，与目前的药物形式相比具有优势，尤其是那些容易产生耐药性的小分子药物。在这里，我们评估了一种噬菌体P22病毒样颗粒（VLP）系统，其中BAPs作为与P22支架蛋白（SP）的融合蛋白被包裹在大肠杆菌中自组装的蛋白笼内。来自三类结构不同的BAPs的代表性肽段以高载量与VLP衣壳蛋白（CP）的比例成功包裹进P22 VLPs中，这对应于SP-BAPs紧密的正电结构与CP亚基向内表面的负电区域之间的相互作用。然而，高负载密度并不对应于提高的SP-BAP产量。这项研究的一个意外发现是，虽然包裹减轻了SP-BAPs对大肠杆菌生长的负面影响，但P22支架蛋白作为积累BAPs的足够融合伙伴，CP的共表达并没有进一步提高SP-BAP产量。尽管如此，在VLPs中包裹为生产功能上等同于其合成对应物的线性和环状重组（r）BAPs的纯化流程提供了有用的第一步。需要进一步努力优化CP与SP-BAP融合体的表达比例，以充分发挥包裹在保护表达宿主和最大化rBAP产量方面的潜力。
