Cell wall and autolytic system of Lactobacillus helveticus ATCC 12046

Abstract

The cell wall of Lactobacillus helveticus was found to have a three-layered structure when thin sections of whole cells or isolated cell walls were stained by various procedures and examined by electron microscopy. The main feature of the composition of isolated cell walls was a high protein content (about 48% of the cell wall dry weight) with a predominant 52 kDa protein. Peptidoglycan and what was considered as teichoic acids accounted for 36% and 14%, respectively, of the cell wall dry weight. The 52 kDa protein was apparently unglycosylated, located in the outer layer and noncovalently bound to the inner layers. Peptidoglycan was mostly assigned to the innermost layer. The conditions leading to autolysis of whole cells or isolated cell walls were determined, as well as the specificity of the autolysin involved in cell wall degradation. IntroductionLactobacillus helveticus is an important lactic acid bacterium that is involved in cheese technology (Accolas et al., 1980;Turner et al., 1983) and that has recently become the object of interest for genetic manipulation (Chassy, 1987;Sandine, 1987;Thomas & Collins, 1988). Its autolysis with the concomitant release of cytoplasmic proteases (Ezzat et al., 1986) is an essential feature of cheese maturation (Bie & Sjostrom, 1975a, b). A better understanding of the autolytic system of L. helveticus could perhaps offer a new way for an accelerated ripening of cheese. Ohmiya & Sat0 (1970) were the first to follow the autolysis of L. helveticus in technological conditions. After adding whole cells to an aseptic rennet curd, the release of DNA was noted and autolysed cells were clearly observed by electron microscopy. More recently, we promoted the autolysis of L. helveticus by suspending whole cells in dilute buffers (Lortal et al., 1989). A clear understanding of the autolytic system of L. helveticus implies, in particular, a better knowledge of the composition and structure of its cell wall, which have so far been investigated only to a limited extent (Ikawa & Snell, 1960;Baddiley & Davison, 1961 ; Williams, 1971). The main peptidoglycan amino acids were determined (Glu, Ala, Asp, Lys) as well as their respective percentage in the D-configuration (Ikawa & Snell, 1960). Previous studies also revealed the presence of glyceroltype teichoic acids (Baddiley & Davison, 1961) and neutral sugars (glucose, galactose but no rhamnose). Wall and membrane teichoic acids of L. helveticus were shown to have antigenic properties (Knox & Wicken, 1971). It has also been suggested that the cell wall of L. helveticus ATCC 10797 was composed of two layers with the outer layer presumably containing a predominant protein (Masuda & Kawata, 1983).In the work described here, a more detailed investigation of the morphology and composition of the cell wall of L. helveticus ATCC 12046 was undertaken. Moreover, the main characteristics of the autolytic system of this strain were investigated, in particular ways of promoting lysis, the effect of external factors, and the specificity of the autolytic...

Microbiological characteristics of Parmigiano Reggiano cheese during the cheesemaking and the first months of the ripening

“…However, concurring with previous publications, the thermophilic bacteria generally disappeared in a brief time (1-3 months), thus releasing a wealth of enzymes [3,7,8,17,31]. An undoubtedly important contribution was made by the mesophilic lactic acid bacteria (essentially L. casei, L. paracasei, L. rhamnosus, pediococci) which presented meaningful counts in the fifth month of ageing, thereby showing a significantly higher level compared to what has been reported thus far in literature with specific reference to Grana Padano [1].…”

X-ray photoelectron spectroscopy and biochemical analysis of the surface of Lactobacillus helveticus ATCC 12046

“…Quantitative amino acid and hexosamine analyses were performed as described by Lortal et al (1991). The phosphorus content was determined according to Eckman (1993) using sodium dihydrogen phosphate as standard.…”

“…LiCl treatment eliminates the outer layer observed by electron microscopy, so the surface exposed after this treatment is considered to be the intermediate layer (Lortal et al, 1991). Given the high surface oxygen and low surface nitrogen of treated cells (Table 2), the high proportion of the carbon peak component at 286.3 eV (Table 3) )I Hydrocarbon-like compound (CH,).…”

S-layer of Lactobacillus helveticus ATCC 12046: isolation, chemical characterization and re-formation after extraction with lithium chloride

“…Previous work (Lortal et al, 1991) showed that the outermost layer of the cell wall was composed of a nonglycosylated 52 kDa protein. In this investigation, several procedures for extracting the S-layer protein from whole cells and isolated cell walls were tested (Table 1).…”

“…In previous work (Lortal et at., 1991) the cell wall of L. helveticus ATCC 12046 was found to have a three-layered structure. The external layer was shown to be composed of a 52 kDa protein, non-covalently linked to the internal layers.…”