HEBERT Elvira Maria
congresos y reuniones científicas
Proteinase activity of Streptococcus thermophilus strains belonging to CRL culture collection.
Simposio; IV International Symposium on Lactic Acid Bacteria: Food, Health and Applications.; 2013
The lactic acid bacterium (LAB) Streptococcus thermophilus is industrially used for the manufacture of yogurt and several cheeses. To apply a rational use of streptococci strains, the culture collection of CERELA (CRL) has performed a systematic study on different features of Str. thermophilus. This work addresses the proteinase activity (PrtS) of a large number (n=52) of Str. thermophilus strains since this property ensures a good behaviour of Str thermophilus in milk and its application as dairy starter. The PrtS activity of Str. thermophilus CRL strains grown in a chemically defined medium (CDM), was evaluated with the chromogenic substrate succinyl-alanil-alanil-prolil-phenylalanine-pnitroanilide (S-ALA). Cell growth and acidifying capacity were evaluated in 10% (w/v) reconstituted skim milk (RSM). In PrtS+ strains, proteolysis of B-casein was analyzed by SDS-PAGE and released amino acid by the o-phthaldialdehyde (o-PA) method. . Five out of 52 strains (9%) showed PrtS activity (PrtS+ strains), fact that was coincident with high growth (µ = 1.25 h-1 ± 0.03; 9.20 ± 0.14 Log CFU/ml) and acidification (pH 4.80 - 4.96 at 4h) in RSM. The multivariate principal component analysis grouped these strains separately from those PrtS- (n = 47), showing that PrtS activity is mainly correlated with the acidifying capacity of the strains. The PrtS+ strains revealed two interesting phenotypes (p<0.05); higher acidifying capacity in RSM (phenotype A, strains CRL1597-1598) and PrtS activity in CDM (phenotype B, strains CRL1764-1765-1767), respectively. Nosignificant differences (p>0.05) were observed in RSM growth and in amino acid released from B-casein (1.08 ± 0.10 µmoles/ml), while the SDS-PAGE analysis showed higher PrtS activity in strains CRL1764-1767. To characterize the PrtS+ strains, RAPD-PCR assays were performed with primers M13, XD9, RAPD1 and RAPD2. Two profiles were found, corresponding to strains with phenotype A and B described previouslty. Str. thermophilus CRL1598 was selected to characterize PrtS activity in CDM at free and controlled pH (6.50) and at different phases of growth. At controlled pH fermentations, maximum cell viability (3.6 x 109 CFU/ml), lactic acid production (17.7 g/l) and PrtS activity (30.92 EU/Log CFU) were obtained as compared to free pH cultures. It is worth to notice that PrtS activity increased continuously in cultures at pH 6.50 while at free pH fermentation, after reaching the maximum at 2h, became undetectable at 6 h. The presented results showed that the cell-envelope proteinase PrtS is a key determinant of the behaviour in milk in Str. thermophilus, being PrtS correlated with the rate of milk acidification, an issue of major technological importance for dairy industry.