Cloning and Characterization of Two Lactobacillus casei Genes Encoding a Cystathionine Lyase

Abstract

Volatile sulfur compounds are key flavor compounds in several cheese types. To better understand the metabolism of sulfur-containing amino acids, which certainly plays a key role in the release of volatile sulfur compounds, we searched the genome database of Lactobacillus casei ATCC 334 for genes encoding putative homologs of enzymes known to degrade cysteine, cystathionine, and methionine. The search revealed that L. casei possesses two genes that putatively encode a cystathionine ␤-lyase (CBL; EC 4.4.1.8). The enzyme has been implicated in the degradation of not only cystathionine but also cysteine and methionine. Recombinant CBL proteins catalyzed the degradation of L-cystathionine, O-succinyl-L-homoserine, L-cysteine, L-serine, and L-methionine to form ␣-keto acid, hydrogen sulfide, or methanethiol. The two enzymes showed notable differences in substrate specificity and pH optimum.Volatile sulfur compounds (VSC) are found in many cheese varieties and are important components of flavor (10). The sulfur compounds methanethiol, methional, and dimethyl trisulfide (DMTS) are key odorants of Swiss Gruyère cheese flavor and are presumably derived from the bacterial metabolism of methionine, which is present in caseins at higher concentrations than cysteine. Swiss Gruyère (place of designated origin) cheese is produced with thermophilic mixed-strain starters which usually contain Streptococcus thermophilus and Lactobacillus delbrueckii subsp. lactis. In addition, depending on the cheese factory, Lactobacillus helveticus can also be present. However, after ripening, Lactobacillus casei and Lactobacillus rhamnosus bacteria are found in high numbers (8). Interestingly, L. casei is also a constituent of the nonstarter lactic acid bacteria of many other cheese varieties (6). Therefore, this species is considered to play an important role during ripening and may be a key player in the formation of flavor compounds.The catabolic pathway(s) of methionine and cysteine in lactobacilli is not well characterized. Possible pathways involve transaminases, methionine ␥-lyases, cystathionine ␤-lyases (CBL), and cystathionine ␥-lyases (CGL) (13). The transamination of methionine by aminotransferases has been studied in lactococci (18,27,34). The results of these studies suggested that methionine is converted to 2-oxo-4-methylthiobutyric acid, which is then converted, either enzymatically or chemically, to methanethiol. In Brevibacterium linens, a methionine ␥-lyases which catalyzes the deamination of methionine and releases methanethiol has been identified and the corresponding gene has been cloned (2,14).CBL, which is encoded by the gene metC in Escherichia coli, is involved in the ␣,␤-elimination of cystathionine to form homocysteine, pyruvate, and ammonia. In comparison, CGL catalyzes the ␣,␥-elimination reaction of cystathionine, producing cysteine, ammonia, and ␣-ketobutyrate. CBL and CGL generally show catalytic promiscuity, which means that they catalyze different reactions. Thus, the CBL from Lactococcus lactis subsp. cremor...

Cystathionine γ-Lyase Is a Component of Cystine-Mediated Oxidative Defense in Lactobacillus reuteri BR11

“…MetCtype cystathionine lyases of Lactobacillus spp. have been reported to occur in Lactobacillus fermentum DT41 (43), Lactobacillus helveticus CNRZ 32 (28), and Lactobacillus casei FAM18168 (20). Sequences are available for only the last two, and they have identities of 65% and 45% and similarities of 81% and 67%, respectively, to Cgl of BR11 at the amino acid level.…”

“…Cystathionine lyases of LAB have been widely studied in the context of cheese flavor development (4,5,20,28,43). However, there are few studies about the physiological roles of cystathionine lyases of LAB.…”

In Helicobacter pylori , LuxS Is a Key Enzyme in Cysteine Provision through a Reverse Transsulfuration Pathway

“…Thus, the true physiological role of MetB Hp remained unclear. With regard to CysK Hp , a similar enzyme from Lactobacillus casei, also annotated CysK, has recently been proposed to act as a cystathionine ␤-synthase, converting homocysteine to cystathionine (17). CysK Hp has up to now been thought to act as a cysteine synthase that uses sulfide to generate cysteine (10).…”

Screening, morphological and molecular characterization of fungi producing cystathionine γ-lyase

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