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Recombinant wheat endoplasmic reticulum oxidoreductin 1 (wEro1) with considerable ability was expressed in Escherichia coli. The functional roles of wEro1 in flour processing quality were investigated by farinographic, rheological, texture profile analysis, electrophoresis, size exclusion chromatography, scanning electron microscopy, and Fourier transform infrared spectroscopy. wEro1 exhibited an obvious oxidation activity of sulfhydryl groups in small molecule and protein. Addition of wEro1 could strengthen the processing quality of dough, indicated by the improved mixing characteristics, viscoelastic properties, and bread qualities. These improvement effects of wEro1 could be attributed to the formation of macromolecular gluten polymers and massive gluten networks by disulfide cross-linking. Additionally, the increased β-turn structure further demonstrated the enhancement of dough strength. Moreover, the amount of peroxide in dough was improved significantly from 2.36 to 2.82 μmol/g of flour with 0.15% wEro1 treatment. Therefore, the results suggested that wEro1 is a promising novel flour improver.