Adhesion to cellulose of the Gram-positive bacterium Ruminococcus albus involves type IV pili.
This study was aimed at characterizing a cell-surface 25 kDa glycoprotein (GP25) that was previously shown to be underproduced by a spontaneous adhesion-defective mutant D5 of Ruminococcus albus 20. An antiserum against wild-type strain 20 was adsorbed with the mutant D5 to enrich it in antibodies 'specific' to adhesion structures of R. albus 20. The resulting antiserum, called anti-Adh serum, blocked adhesion of R. albus 20 and reacted mainly with GP25 in bacterial and extracellular protein fractions of R. albus 20. The N-terminal sequence of purified GP25 was identical to that of CbpC, a 21 kDa cellulose-binding protein (CBP) of R. albus 8. The nucleotide sequence of the gp25 gene was determined by PCR and genomic walking procedures. The gp25 gene encoded a protein of 165 aa with a calculated molecular mass of 16940 Da that showed 72.9% identity with CbpC and presented homologies with type IV pilins of Gram-negative pathogenic bacteria. Negative-staining electron microscopy revealed fine and flexible pili surrounding R. albus 20 cells while mutant cells were not piliated. In addition, immunoelectron microscopy showed that the anti-Adh serum probing mainly GP25, completely decorated the pili surrounding R. albus 20, thereby showing that GP25 was a major pilus subunit. This study shows for the first time the presence of pili at the surface of R. albus and identifies GP25 as their major protein subunit. Though GP25 was not identified as a CBP, isolated pili were shown to bind cellulose. In conclusion, these pili, which belong to the family of type IV pili, mediate adhesion of R. albus 20 to cellulose.
Rakotoarivonina
Harivony
H
Unité de Microbiologie1 and Unité de Recherches sur la Viande, Equipe Microbiologie2, INRA, Centre de Recherches de Clermont-Ferrand-Theix, 63122 Saint-Genès-Champanelle, France.
Jubelin
Grégory
G
Unité de Microbiologie1 and Unité de Recherches sur la Viande, Equipe Microbiologie2, INRA, Centre de Recherches de Clermont-Ferrand-Theix, 63122 Saint-Genès-Champanelle, France.
Hebraud
Michel
M
Unité de Microbiologie1 and Unité de Recherches sur la Viande, Equipe Microbiologie2, INRA, Centre de Recherches de Clermont-Ferrand-Theix, 63122 Saint-Genès-Champanelle, France.
Gaillard-Martinie
Brigitte
B
Unité de Microbiologie1 and Unité de Recherches sur la Viande, Equipe Microbiologie2, INRA, Centre de Recherches de Clermont-Ferrand-Theix, 63122 Saint-Genès-Champanelle, France.
Forano
Evelyne
E
Unité de Microbiologie1 and Unité de Recherches sur la Viande, Equipe Microbiologie2, INRA, Centre de Recherches de Clermont-Ferrand-Theix, 63122 Saint-Genès-Champanelle, France.
Mosoni
Pascale
P
Unité de Microbiologie1 and Unité de Recherches sur la Viande, Equipe Microbiologie2, INRA, Centre de Recherches de Clermont-Ferrand-Theix, 63122 Saint-Genès-Champanelle, France.
D000595:Amino Acid Sequence; D000907:Antibodies, Bacterial / Q000276:immunology / Q000494:pharmacology; D001422:Bacterial Adhesion / Q000187:drug effects; D015153:Blotting, Western; D002482:Cellulose / Q000378:metabolism*; D010861:Fimbriae, Bacterial / Q000276:immunology / Q000378:metabolism*; D006023:Glycoproteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D006094:Gram-Positive Bacteria / Q000166:cytology* / Q000276:immunology / Q000378:metabolism* / Q000648:ultrastructure; D008969:Molecular Sequence Data
D000907:Antibodies, Bacterial; D006023:Glycoproteins; D002482:Cellulose