A novel single-chain enzyme complex with chain reaction properties rapidly producing thromboxane A2 and exhibiting powerful anti-bleeding functions

Abstract

Uncontrollable bleeding is still a worldwide killer. In this study, we aimed to investigate a novel approach to exhibit effective haemostatic properties, which could possibly save lives in various bleeding emergencies. According to the structure-based enzymatic design, we have engineered a novel single-chain hybrid enzyme complex (SCHEC), COX-1-10aa-TXAS. We linked the C-terminus of cyclooxygenase-1 (COX-1) to the N-terminus of the thromboxane A₂ (TXA₂ ) synthase (TXAS), through a 10-amino acid residue linker. This recombinant COX-1-10aa-TXAS can effectively pass COX-1-derived intermediate prostaglandin (PG) H₂ (PGH₂ ) to the active site of TXAS, resulting in an effective chain reaction property to produce the haemostatic prostanoid, TXA₂ , rapidly. Advantageously, COX-1-10aa-TXAS constrains the production of other pro-bleeding prostanoids, such as prostacyclin (PGI₂ ) and prostaglandin E₂ (PGE₂ ), through reducing the common substrate, PGH₂ being passed to synthases which produce aforementioned prostanoids. Therefore, based on these multiple properties, this novel COX-1-10aa-TXAS indicated a powerful anti-bleeding ability, which could be used to treat a variety of bleeding situations and could even be useful for bleeding prone situations, including nonsteroidal anti-inflammatory drugs (NSAIDs)-resulted TXA₂ -deficient and PGI₂ -mediated bleeding disorders. This novel SCHEC has a great potential to be developed into a biological haemostatic agent to treat severe haemorrhage emergencies, which will prevent the complications of blood loss and save lives.

Keywords: enzyme engineering; haemostasis; prostanoids; thromboxane A2.