Conserved residues in the CHASE3 domain of BmsA are involved in motility and biofilm formation in Pseudomonas alkylphenolica KL28

Abstract The bmsA gene of Pseudomonas alkylphenolica KL28T encodes a conserved protein involved in motility and multicellular biofilm formation. Sequence analysis revealed that BmsA contains an extracellular CHASE3 sensory domain, suggesting a role in environmental signal perception. To investigate its function, a CHASE3 domain deletion mutant and six alanine substitution mutants targeting conserved residues, including Arg73 and Phe75 within the RG(Y/F) motif, were constructed and introduced into a bmsA mutant background. SDS–PAGE confirmed the production of BmsA and its variants, although most proteins migrated at approximately 145 kDa, above the predicted 130.3 kDa. Phenotypic and motility analyses showed that deletion of the CHASE3 domain and substitution of Arg73 resulted in smooth colony morphology and reduced pellicle and aerial structure formation, similar to the phenotypes observed in the bmsA mutant. In addition, the Arg73 substitution exhibited increased swimming motility in soft agar, resembling the bmsA mutant phenotype. The other alanine substitutions showed no substantial phenotypic differences compared with the control strain. These results indicate that the conserved Arg73 residue within the RG(Y/F) motif of the CHASE3 domain contributes to the modulation of BmsA-dependent phenotypes in strain KL28.