The plant pathogen Pseudomonas syringae accounts for substantial crop losses and is considered an important agricultural issue. To better manage disease in the field, it is important to have an understanding of the underlying genetic mechanisms that mediate virulence. There are a substantial number of genes in sequenced bacterial genomes, including P. syringae, that encode for conserved hypothetical proteins; some of these have been functionally characterized in other Pseudomonads and have been demonstrated to play important roles in disease. PSPTO_3957 encodes a conserved hypothetical protein of unknown function. To evaluate the role of PSPTO_3957 in P. syringae pv. tomato DC3000, a PSPTO_3957 deletion mutant was constructed. Here, we show that PSPTO_3957 does not influence growth on rich media, motility or biofilm formation but is necessary for nitrate assimilation and full virulence in P. syringae. Our results have revealed an important role for PSPTO_3957 in the biology of P. syringae. Given the conservation of this protein among many bacteria, this protein might serve as an attractive target for disease management of this and other bacterial plant pathogens.