Thioredoxins comprise a growing family of proteins that function as general protein-disulfide reductases and are maintained in their reduced active form by the flavoenzyme thioredoxin reductase. Human Trx-1 is mainly a cytosolic protein, although it has been shown to translocate into the nucleus upon certain stimuli and can also be secreted. We report here the expression and characterization of Delta3Trx-1, a splicing variant of human Trx-1, lacking exon 3, which spans from residues 44 to 63 in the wild-type protein. Structure-based prediction of this splicing form indicates that A3Trx-1 lacks helix alpha2 and strand beta3, which are implicated in substrate positioning and three-dimensional stabilization of the active site residues. Recombinant human Delta3Trx-1 is recognized by polyclonal antibodies raised against full-length human Trx-1. However, Delta3Trx-1 retains no enzymatic activity either with DTT or thioredoxin reductase and NADPH as reducing systems. Delta3Trx-1 competes with full-length Trx-1 for the interaction with thioredoxin reductase. The absence of helix alpha2 and strand beta3 in A3Trx-1 is consistent with the lack of enzymatic activity and its potential dominant negative properties. (C) 2002 Elsevier Science (USA). All rights reserved.