Mhp107 Is a Member of the Multifunctional Adhesin Family of Mycoplasma hyopneumoniae

Abstract

Mycoplasma hyopneumoniae is the causative pathogen of porcine enzootic pneumonia, an economically significant disease that disrupts the mucociliary escalator in the swine respiratory tract. Expression of Mhp107, a P97 paralog encoded by the gene mhp107, was confirmed using ESI-MS/MS. To investigate the function of Mhp107, three recombinant proteins,

Sequence TTKF↓QE Defines the Site of Proteolytic Cleavage in Mhp683 Protein, a Novel Glycosaminoglycan and Cilium Adhesin of Mycoplasma hyopneumoniae

“…We have previously established that proteolytic processing of members of the P97 and P102 paralog families generates a complex array of cleavage fragments that are displayed on the surface of M. hyopneumoniae and perform key roles in adhesion to structurally diverse host molecules (13)(14)(15)(16)(17)(18)48). However, the rationale for the extensive proteolytic processing of these 3 M. P. Padula and S. P. Djordjevic, unpublished data.…”

“…The solution was removed from the wells, and the gel pieces were vacuum-dried for 1 h. The gel spots were rehydrated in 8 l of trypsin solution (2 ng l Ϫ1 (sequencing-grade modified trypsin (Promega, Madison WI) in 40 mM ammonium bicarbonate)) at 4°C for 1 h. Excess trypsin was removed, and the gel pieces were resuspended in 25 l of 40 mM ammonium bicarbonate and incubated overnight at 37°C. Peptides were concentrated and desalted using C 18 Perfect Pure TM tips (Eppendorf, Hamburg, Germany) and eluted in matrix (␣-cyano-4-hydroxycinnamic acid (Sigma), 8 mg ml Ϫ1 in 70% (v/v) acetonitrile, 1% (v/v) formic acid) directly onto a target plate. Peptide mass maps were generated by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) using a Voyager DE-STR (Applied Biosystems, Framingham MA).…”

“…Mhp683 Binds Heparin-In previous studies we have identified members of the P97 family as heparin-binding proteins (15,16,18,48). To determine whether Mhp683 binds glycosaminoglycans, we performed microtiter plate assays with biotin-labeled heparin.…”

“…Tandem pentapeptide repeats (AAKP(V/E)) in the R1 domain of P97 (19,20) play an important role in cilium binding. Non-R1-containing members of the P97 family also bind porcine cilia (15,18) and epithelial cell surfaces (13,18). The P97 paralog Mhp107 also binds plasminogen, fibronectin, and the glycosaminoglycan heparin (18).…”

“…Non-R1-containing members of the P97 family also bind porcine cilia (15,18) and epithelial cell surfaces (13,18). The P97 paralog Mhp107 also binds plasminogen, fibronectin, and the glycosaminoglycan heparin (18). Mhp108, a member of the P102 family, binds cilia, fibronectin, and plasminogen (17), and domains within Mhp183, Mhp494, Mhp493, and Mhp271 bind heparin.…”