Hasselt University, Biomedical Research Institute, Agoralaan, Diepenbeek, Belgium.
Proteomics. 2011 Jun;11(12):2550-4. doi: 10.1002/pmic.201000783. Epub 2011 May 16.
Phage display is a high-throughput technology used to identify ligands for a given target. A drawback of the approach is the absence of PTMs in phage-displayed peptides. The applicability of phage display could be broadened considerably by the implementation of PTMs in this system. The aim of this study was to investigate the possible application of citrullination, a PTM of an arginine into a citrulline amino acid, in filamentous (M13) and lytic (T7) phage display. After in vitro citrullination of T7 and M13 phages, citrullination was confirmed and the infectivity of both citrullinated and non-citrullinated phage was compared by titer determination. We demonstrated the successful in vitro citrullination of T7 and M13 phage-displayed peptides. This in vitro modification did not affect the viability or infectivity of the T7 virions, a necessary prerequisite for the implementation of this approach in T7 phage display. For M13 phage, however, the infecting phage titer decreased five-fold upon citrullination, limiting the use of this modification in M13 phage display. In conclusion, in vitro citrullination can be applied in T7 phage display giving rise to a high-throughput and sensitive approach to identify citrulline-containing ligands by the use of the strengths of phage display technology.
噬菌体展示是一种高通量技术，用于鉴定给定靶标的配体。该方法的一个缺点是噬菌体展示的肽中缺乏 PTMs。通过在该系统中实施 PTMs，可以大大拓宽噬菌体展示的适用性。本研究的目的是研究精氨酸的一种 PTM 即瓜氨酸化，在丝状（M13）和裂解（T7）噬菌体展示中的可能应用。在体外对 T7 和 M13 噬菌体进行瓜氨酸化后，通过滴定确定了瓜氨酸化的确认和感染性，比较了瓜氨酸化和非瓜氨酸化噬菌体的感染性。我们证明了 T7 和 M13 噬菌体展示肽的体外瓜氨酸化成功。这种体外修饰不影响 T7 病毒粒子的活力或感染性，这是在 T7 噬菌体展示中实施这种方法的必要前提。然而，对于 M13 噬菌体，瓜氨酸化后感染噬菌体的滴度降低了五倍，限制了这种修饰在 M13 噬菌体展示中的应用。总之，体外瓜氨酸化可应用于 T7 噬菌体展示，通过利用噬菌体展示技术的优势，提供一种高通量和敏感的方法来鉴定含瓜氨酸的配体。
