Identification of Mfa5-2 as a structural component of Mfa1 fimbriae in Porphyromonas gingivalis

OBJECTIVES: The periodontal pathogen Porphyromonas gingivalis produces Mfa1 fimbriae, which play an important role in biofilm formation and bacterial colonization. In P. gingivalis ATCC 33277, Mfa1 fimbriae are composed of the major Mfa1 subunit and the accessory proteins Mfa3, Mfa4, and Mfa5. Some P. gingivalis strains harbor an additional Mfa5-related gene (mfa5-2). However, mfa5-2 presence in the genome of P. gingivalis EM3 and 1439 and Mfa5-2 incorporation into the Mfa1 fimbriae in both strains remain unclear. Therefore, this study investigated mfa5-2 presence in the genomes of the two bacterial strains and assessed Mfa5-2 incorporation into the Mfa1 fimbriae. METHODS: The genomic regions downstream of mfa5 in P. gingivalis EM3 and 1439 were sequenced. Subsequently, Mfa1 fimbriae were purified and analyzed using sodium dodecyl-sulfate polyacrylamide gel electrophoresis, followed by mass spectrometry and immunoblotting with an Mfa5-2-specific peptide antibody. Fimbrial morphology was examined using transmission electron microscopy. RESULTS: Sequencing results revealed that mfa5-2 was located immediately downstream of mfa5-1 in both strains. The purified fimbrial fractions contained Mfa1, Mfa3, Mfa4, Mfa5, and a protein of approximately 250 kDa, identified as Mfa5-2. Transmission electron microscopy showed that fimbrial architecture remained unchanged despite Mfa5-2 incorporation. CONCLUSIONS: Mfa5-2 is expressed as a high-molecular-weight component and integrated into the Mfa1 fimbriae in P. gingivalis EM3 and 1439. These findings revealed previously unknown structural diversity of Mfa1 fimbriae.