prtH2, not prtH, is the ubiquitous cell wall proteinase gene in Lactobacillus helveticus.
Lactobacillus helveticus strains possess an efficient proteolytic system that releases peptides which are essential for lactobacillus growth in various fermented dairy products and also affect textural properties or biological activities. Cell envelope proteinases (CEPs) are bacterial enzymes that hydrolyze milk proteins. In the case of L. helveticus, two CEPs with low percentages of amino acid identity have been described, i.e., PrtH and PrtH2. However, the distribution of the genes that encode CEPs still remains unclear, rendering it difficult to further control the formation of particular peptides. This study evaluated the diversity of genes that encode CEPs in a collection of strains of L. helveticus isolated from various biotopes, both in terms of the presence or absence of these genes and in terms of nucleotide sequence, and studied their transcription in dairy matrices. After defining three sets of primers for both the prtH and prtH2 genes, we studied the distribution of the genes by using PCR and Southern blotting experiments. The prtH2 gene was ubiquitous in the 29 strains of L. helveticus studied, whereas only 18 of them also exhibited the prtH gene. Sequencing of a 350-bp internal fragment of these genes revealed the existence of intraspecific diversity. Finally, expression of these two CEP-encoding genes was followed during the growth in dairy matrices of two strains, ITG LH77 and CNRZ32, which possess one and two CEP-encoding genes, respectively. Both genes were shown to be expressed by L. helveticus at each stage of growth in milk and at different stages of mini-Swiss-type cheese making and ripening.
Genay
M
M
Unité de Recherche sur l'Animal et les Fonctionnalités des Produits Animaux, U.C. INRA 340, Nancy-Université, Vandoeuvre-lès-Nancy, France.
Sadat
L
L
Gagnaire
V
V
Lortal
S
S
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics* / Q000302:isolation & purification*; D015139:Blotting, Southern; D002473:Cell Wall / Q000201:enzymology*; D002611:Cheese / Q000382:microbiology*; D017931:DNA Primers / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics; D021901:DNA, Intergenic; D010450:Endopeptidases / Q000235:genetics* / Q000302:isolation & purification*; D020869:Gene Expression Profiling; D014644:Genetic Variation; D052198:Lactobacillus helveticus / Q000201:enzymology*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D012689:Sequence Homology, Nucleic Acid
D001426:Bacterial Proteins; D017931:DNA Primers; D004269:DNA, Bacterial; D021901:DNA, Intergenic; D010450:Endopeptidases; C120659:PrtH protein, Lactobacillus