Identification and genetic characterization of a novel proteinase, PrtR, from the human isolate Lactobacillus rhamnosus BGT10.
A novel proteinase, PrtR, produced by the human vaginal isolate Lactobacillus rhamnosus strain BGT10 was identified and genetically characterized. The prtR gene and flanking regions were cloned and sequenced. The deduced amino acid sequence of PrtR shares characteristics that are common for other cell envelope proteinases (CEPs) characterized to date, but in contrast to the other cell surface subtilisin-like serine proteinases, it has a smaller and somewhat different B domain and lacks the helix domain, and the anchor domain has a rare sorting signal sequence. Furthermore, PrtR lacks the insert domain, which otherwise is situated inside the catalytic serine protease domain of all CEPs, and has a different cell wall spacer (W) domain similar to that of the cell surface antigen I and II polypeptides expressed by oral and vaginal streptococci. Moreover, the PrtR W domain exhibits significant sequence homology to the consensus sequence that has been shown to be the hallmark of human intestinal mucin protein. According to its alpha(S1)- and beta-casein cleavage efficacy, PrtR is an efficient proteinase at pH 6.5 and is distributed throughout all L. rhamnosus strains tested. Proteinase extracts of the BGT10 strain obtained with Ca(2+)-free buffer at pH 6.5 were proteolytically active. The prtR promoter-like sequence was determined, and the minimal promoter region was defined by use of prtR-gusA operon fusions. The prtR expression is Casitone dependent, emphasizing that nitrogen depletion elevates its transcription. This is in correlation with the catalytic activity of the PrtR proteinase.
Pastar
Irena
I
Institute of Molecular Genetics and Genetic Engineering, 11001 Belgrade, Serbia and Montenegro. Wageningen Centre for Food Sciences, 6700 AN Wageningen. NIZO Food Research, 6710 BA Ede, The Netherlands.
Tonic
Ivana
I
Golic
Natasa
N
Kojic
Milan
M
van Kranenburg
Richard
R
Kleerebezem
Michiel
M
Topisirovic
Ljubisa
L
Jovanovic
Goran
G
D000595:Amino Acid Sequence; D001426:Bacterial Proteins; D001483:Base Sequence; D002364:Caseins / Q000378:metabolism; D002473:Cell Wall / Q000201:enzymology; D003001:Cloning, Molecular; D003470:Culture Media; D003546:Cysteine Endopeptidases / Q000737:chemistry* / Q000235:genetics* / Q000302:isolation & purification; D005260:Female; D015964:Gene Expression Regulation, Bacterial; D006801:Humans; D007778:Lactobacillus / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D017422:Sequence Analysis, DNA; D014621:Vagina / Q000382:microbiology*
D001426:Bacterial Proteins; D002364:Caseins; D003470:Culture Media; C475729:casein hydrolysate; D003546:Cysteine Endopeptidases; C091975:prtR protein, Porphyromonas gingivalis W50