The conserved histidine 106 of large thioredoxin reductases is likely to have a structural role but not a base catalyst function.

The catalytic activity of selenocysteine-containing thioredoxin reductases can be mimicked by cysteine-variants if the local environment at the C-terminal redox center supports thiol activation. This concept of a linear catalytic site was challenged by structural data suggesting that the invariant residue His106 functions as a base catalyst for the dithiol-disulphide exchange reaction between enzyme and substrate. As reported here, we changed His106 to asparagine, glutamine, and phenylalanine in various C-terminal mutants of Drosophila melanogaster thioredoxin reductase. The catalytic activity dropped considerably, yet pH-profiles did not reveal differences, rendering a function for His106 as a base catalyst unlikely. Interestingly, the phenylalanine-mutants, designed as negative controls were the most active mutants which suggests rather a structural role of His106.

Synthesis of 5-nitro-2-furancarbohydrazides and their cis-diamminedichloroplatinum complexes as bitopic and irreversible human thioredoxin reductase inhibitors.

The human selenoprotein thioredoxin reductase is involved in antioxidant defense and DNA synthesis. As increased thioredoxin reductase levels are associated with drug sensitivity to cisplatin and drug resistance in tumor cells, this enzyme represents a promising target for the development of cytostatic agents. To optimize the potential of the widely used cisplatin to inhibit the human thioredoxin reductase and therefore to overcome cisplatin resistance, we developed and synthesized four cis-diamminedichloroplatinum complexes of the lead 5-nitro-2-furancarbohydrazide 8 selected from high-throughput screening. Detailed kinetics revealed that the isolated fragments, 5-nitro-2-furancarbohydrazide and cisplatin itself, bind with micromolar affinities at two different subsites of the human enzyme. By tethering both fragments four nitrofuran-based cis-diamminedichloroplatinum complexes 13a-c and 20 were synthesized and identified as bi-ligand irreversible inhibitors of the human enzyme with nanomolar affinities. Studies with mutant enzymes clearly demonstrate the penultimate selenocysteine residue as the prime target of the synthesized cis-diamminedichloroplatinum complexes.