Enhanced anti‐tumor activity of arginine decarboxylase through the incorporation of aromatic amino acids at the multimer‐forming interface

The pressing challenge of cancer's high mortality and invasiveness demands improved therapeutic approaches. Targeting the nutrient dependencies within cancer cells has emerged as a promising approach. This study is dedicated to demonstrating the potential of arginine depletion for cancer treatment. Notably, the focus centers on arginine decarboxylase (RDC), a pH‐dependent enzyme expecting enhanced activity within the slightly acidic microenvironments of tumors. To investigate the effect of a single‐site mutation on the catalytic efficacy of RDC, diverse amino acids, including glycine, alanine, phenylalanine, tyrosine, tryptophan, p‐azido‐phenylalanine, and a phenylalanine analog with a hydrogen‐substituted tetrazine, were introduced at the crucial threonine site (position 39) in the multimer‐forming interface. Remarkably, the introduction of either a natural or a non‐natural aromatic amino acid at position 39 substantially boosted enzymatic activity, while amino acids with smaller side chains did not show the same effect. This enhanced enzymatic activity is likely attributed to the reinforced formation of multimer structures through favorable interactions between the introduced aromatic amino acid and the neighboring subunit. Noteworthy, at slightly acidic pH, the RDC variant featuring tryptophan at position 39 demonstrated augmented cytotoxicity against tumor cells compared to the wild‐type RDC. This attribute aligns with the tumor microenvironment and positions these variants as potential candidates for targeted cancer therapy. Graphical and Lay Summary Arginine decarboxylase (RDC) shows promise as a potential cancer drug candidate due to its unique pH‐dependent activity, enabling targeted action within the tumor microenvironment. However, a significant challenge arises from RDC's limited enzymatic activity. In this study, we aim to enhance RDC's activity by introducing an aromatic amino acid substitution at the 39th threonine position. This research provides valuable insights into the development of customized drugs for intricate cellular environments, especially within cancerous tumors.