Determination of the role of recombinant MrkD protein in adhesion to fibroblast cells and collagen

Objective: The present study aimed to investigate, at the molecular level, the interaction of recombinant MrkD protein with human fibroblast cells and type V collagen, and to evaluate its potential as a therapeutic target. Material and Methot: The mrkD gene from K. pneumoniae was amplified using specifically designed primers and cloned into the pET30a(+) expression vector, followed by transformation into E. coli BL21 cells. The recombinant MrkD protein was purified using IMAC. Its cytotoxicity on BJ human fibroblast cells was assessed via the WST-1 assay, and its impact on bacterial adhesion capacity was determined. In addition, ELISA assays were employed to evaluate the interaction between MrkD and type V collagen. Results: The molecular weight of the purified recombinant MrkD protein was determined to be approximately 40.8 kDa. ELISA analyses confirmed the specific interaction between MrkD and type V collagen. WST-1 assay results indicated that MrkD exhibited no cytotoxic effects on BJ fibroblast cells. Furthermore, the presence of recombinant MrkD protein was associated with a reduction of approximately 0.85 log in the adhesion of K. pneumoniae to fibroblast cells. Conclusion: This study provides the first experimental evidence demonstrating the specific interaction between recombinant MrkD protein and human fibroblast cells. It also directly evaluates the cytotoxic potential of MrkD. The findings offer new insights into the collagen-binding capacity of this adhesin and suggest that MrkD may serve as a promising target for anti-adhesion therapeutic strategies against K. pneumoniae infections.