Altered expression and modification of proteases from an avirulent mutant of Porphyromonas gingivalis W50 (W50/BE1).
Proteases of Porphyromonas gingivalis are considered to be important factors in the virulence of this organism. A non-pigmenting mutant of P. gingivalis W50 (W50/BE1) has been shown to be less virulent in animal models and to produce significantly less Arg-specific protease activity than the parent strain. Three proteases are present in the culture supernatant of P. gingivalis W50: RI, RIA and RIB. All three proteases are derived from prpR1, which encodes a polypeptide of 1706 amino acids that is organized into distinct domains (pro, alpha, beta and gamma). The aim of the present investigation was to purify and characterize the Arg-specific proteases produced by the avirulent W50/BE1 strain. Significant differences were observed between the proteases of P. gingivalis W50 and W50/BE1. The levels of RI present in the culture supernatant of W50/BE1 were lower than those present in W50, and RIA and RIB were absent. RI from W50/BE1 was composed of three polypeptide chains, unlike the enzyme from W50, which is a heterodimer. The remainder of the Arg-specific protease activity in W50/BE1 was derived from a second gene, prR2, and was present in two fractions, RIIAs/BE (soluble) and RIIAv/BE (vesicle-bound). This activity contained two peptide chains: a approximately 54 kDa chain corresponding to the protease domain and a approximately 26 kDa chain, derived from the propeptide domain of the PrRII precursor. No enzyme with large glycan additions, equivalent to RIB in the vesicle fraction of the wild-type W50, was present. These data indicate that the reduced level of extracellular protease activity in W50/BE1 reflects reduced synthesis and/or export of prpR1 enzymes, which is only partially compensated by synthesis of prR2-derived enzymes, and that all of these proteases undergo altered post-translational modification compared to the parent strain.
MRC Molecular Pathogenesis Group, Department of Oral Microbiology, St Bartholomew's and the Royal London School of Medicine and Dentistry, Queen Mary and Westfield College, University of London,32 Newark Street, London E1 2AA,UK.
Lucy M
LM
Collinson
MRC Molecular Pathogenesis Group, Department of Oral Microbiology, St Bartholomew's and the Royal London School of Medicine and Dentistry, Queen Mary and Westfield College, University of London,32 Newark Street, London E1 2AA,UK.
Minnie
M
Rangarajan
MRC Molecular Pathogenesis Group, Department of Oral Microbiology, St Bartholomew's and the Royal London School of Medicine and Dentistry, Queen Mary and Westfield College, University of London,32 Newark Street, London E1 2AA,UK.
Michael A
MA
Curtis
D000595:Amino Acid Sequence; D000818:Animals; D000907:Antibodies, Bacterial; D001120:Arginine / Q000378:metabolism; D001426:Bacterial Proteins / Q000276:immunology / Q000302:isolation & purification; D010450:Endopeptidases / Q000235:genetics* / Q000276:immunology / Q000378:metabolism*; D015870:Gene Expression; D005798:Genes, Bacterial; D007700:Kinetics; D008969:Molecular Sequence Data; D009154:Mutation; D016966:Porphyromonas gingivalis / Q000201:enzymology* / Q000235:genetics / Q000472:pathogenicity*; D011499:Protein Processing, Post-Translational; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D014774:Virulence / Q000235:genetics / Q000502:physiology
D000907:Antibodies, Bacterial; D001426:Bacterial Proteins; D001120:Arginine; D010450:Endopeptidases