Novel Extracellular x-Prolyl Dipeptidyl-Peptidase (DPP) from Streptococcus gordonii FSS2: an Emerging Subfamily of Viridans Streptococcal x-Prolyl DPPs

Abstract

Streptococcus gordonii is generally considered a benign inhabitant of the oral microflora, and yet it is a primary etiological agent in the development of subacute bacterial endocarditis (SBE), an inflammatory state that propagates thrombus formation and tissue damage on the surface of heart valves. Strain FSS2 produced several extracellular aminopeptidase and fibrinogen-degrading activities during growth in culture. In this report we describe the purification, characterization, and cloning of a serine class dipeptidyl-aminopeptidase, an x-prolyl dipeptidyl-peptidase (Sg-xPDPP, for S. gordonii x-prolyl dipeptidyl-peptidase), produced in a pH-controlled batch culture. Purification of this enzyme by anion exchange, gel filtration, and hydrophobic interaction chromatography yielded a protein monomer of approximately 85 kDa, as shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (PAGE) under denaturing conditions. However, under native conditions, the protein appeared to be a homodimer on the basis of gel filtration and PAGE. Kinetic studies indicated that purified enzyme had a unique and stringent x-prolyl specificity that is comparable to both the dipeptidyl-peptidase IV/CD26 and lactococcal x-prolyl dipeptidyl-peptidase families. Nested PCR cloning from an S. gordonii library enabled the isolation and sequence analysis of the full-length gene. A 759-amino-acid polypeptide with a theoretical molecular mass of 87,115 Da and a calculated pI of 5.6 was encoded by this open reading frame. Significant homology was found with the PepX gene family from Lactobacillus and Lactococcus spp. and putative x-prolyl dipeptidyl-peptidases from other streptococcal species. Sg-xPDPP may serve as a critical factor for the sustained bacterial growth in vivo and furthermore may aid in the proteolysis of host tissue that is commonly observed during SBE pathology.

Cloning, Purification, and Enzymatic Properties of Dipeptidyl Peptidase IV from the Swine Pathogen Streptococcus suis

“…To a lesser extent, other dipeptides, including Lys-Ala-pNa, Arg-Pro-pNa, Ala-Ala-pNa, and Asp-Pro-pNa, were also hydrolyzed by the protease. A similar observation was previously reported for other bacterial DPP IVs (2,7,17,25).…”

“…The deduced amino acid sequence from the nucleotide sequence revealed that it contained a complete open reading frame coding for a 755-aminoacid protein with a calculated molecular mass of 85 kDa. This correlates well with the molecular masses previously reported for other DPP IVs, including those from P. albensis (27), S. gordonii (7), Lactobacillus sakei (20), and S. thermophilus (25).…”

Contribution of Dipeptidyl Peptidase IV to the Severity of Dextran Sulfate Sodium-Induced Colitis in the Early Phase

“…21) Therefore, short-chain fatty acids can be expected to be a therapeutic alternative for IBD patients by stimulating renewal of the injured epithelium. Homologues of the Dppiv gene have also been found in several bacteria, including Lactobacillus helveticus, 22) Streptococcus gordonii, 23) and Porphyromonas gingivalis. 24) Porphyromonas gingivalis invades connective tissue and attaches to the extracellular matrix because of the interaction between fibronectin and DPPIV.…”

StreptococcusAdherence and Colonization

“…Some of these carry the LPxTz motif for covalent cell wall anchorage, and some do not and are released into the environment (Tables 1 and 2). A dipeptidase from S. gordonii is secreted and released but carries no signal peptide (195). Clearly, there is still a limited understanding of the molecular mechanisms of protein secretion in streptococci.…”