Proteins complexed to the P1 adhesin of Mycoplasma pneumoniae.
Adherence of Mycoplasma pneumoniae to host cells requires several mycoplasmal membrane proteins and cytoskeleton-like proteins in addition to the adhesin P1, a transmembrane protein of 170 kDa. To analyse interactions of the P1 adhesin with other membrane proteins or with cytoskeleton-like proteins, cross-linking studies were performed in vivo using the permeant reagent paraformaldehyde. The cross-linked protein complex was isolated by immunoaffinity chromatography, and proteins complexed to the P1 protein were identified by immunoblot analysis followed by high mass accuracy tryptic peptide mapping using matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS). In addition to the P1 protein and a truncated form of the same protein, the adhesin-related 30 kDa protein, two membrane proteins of 40 and 90 kDa, the cytoskeleton-associated 65 kDa protein and two cytoskeleton-forming proteins, HMW1 and HMW3, were found to be components of the isolated protein complex. Furthermore, the cross-linked complex contained the chaperone DnaK and the E1alpha subunit of pyruvate dehydrogenase. In summary, it was shown that cytadherence-associated membrane proteins are located in close proximity to cytoskeleton-like proteins, suggesting a functional interaction between membrane and cytoskeleton-like proteins. DnaK might be involved in translocation of proteins from the cytoplasm to the membrane and pyruvate dehydrogenase might be a structural protein of the attachment organelle.
Layh-Schmitt
Gerlinde
G
Hygiene-Institut, Universität Heidelberg, Im Neuenheimer Feld 324, 69120 Heidelberg, Germany1.
Podtelejnikov
Alexandre
A
EMBL, Meyerhofstraße 1, 69123 Heidelberg, Germany2.
Mann
Matthias
M
EMBL, Meyerhofstraße 1, 69123 Heidelberg, Germany2.
D018829:Adhesins, Bacterial / Q000378:metabolism*; D001422:Bacterial Adhesion; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism*; D002846:Chromatography, Affinity / Q000379:methods; D003432:Cross-Linking Reagents; D004591:Electrophoresis, Polyacrylamide Gel; D005557:Formaldehyde; D015151:Immunoblotting; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism; D009177:Mycoplasma pneumoniae / Q000254:growth & development / Q000378:metabolism*; D011108:Polymers; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D018829:Adhesins, Bacterial; D001426:Bacterial Proteins; D003432:Cross-Linking Reagents; D008565:Membrane Proteins; D011108:Polymers; C035471:adhesin, Mycoplasma pneumoniae; D005557:Formaldehyde; C003043:paraform