Purification and characterization of a branched-chain amino acid aminotransferase from Lactobacillus paracasei subsp. paracasei CHCC 2115

Abstract

Aim: Purification and characterization of an aminotransferase (AT) specific for the degradation of branched-chain amino acids from Lactobacillus paracasei subsp. paracasei CHCC 2115. Methods and Results:The purification protocol consisted of anion exchange chromatography, affinity chromatography and hydrophobic interaction chromatography. The enzyme was found to exist as a monomer with a molecular mass of 40-50 kDa. The AT converted isoleucine, leucine and valine at a similar rate with a-ketoglutarate as the amino group acceptor; minor activity was shown for methionine. The enzyme had pH and temperature optima of 7AE3 and 43°C, respectively, and activity was detected at the pH and salt conditions found in cheese (pH 5AE2, 4% NaCl). Hg 2+ completely inhibited the enzyme, and the inhibition pattern was similar to that for pyridoxal-5¢-phosphate-dependent enzymes, when studying the effect of other metal ions, thiol-and carbonylbinding agents. The N-terminal sequence of the enzyme was SVNIDWNNLGFDYMQLPYRYVAHXKDGVXD, and had at the amino acid level, 60 and 53% identity to a branched-chain amino acid AT of Lact. plantarum and Lactococcus lactis, respectively. Conclusions: The results suggest that Lact. paracasei subsp. paracasei CHCC 2115 may contribute to development of flavour in cheese. Significance and Impact of the Study: The findings of this work contribute to the knowledge of transamination performed by cheese-related bacteria, and in the understanding and control of amino acid catabolism and the production of aroma compounds.

Potential of three probiotic lactobacilli in transforming star fruit juice into functional beverages

“…α‐Ketoglutaric acid could be formed from the catabolism of glutamic acid (Thage et al., 2004). Oxalic acid remained stable during fermentation (Table 1).…”

“…Isoamyl alcohol could be derived from leucine via amino acid metabolism and is commonly found in foods fermented by Lactobacillus (Thage et al., 2004). Linalool, which gives rise to the citrus and floral aroma in star fruit, was increased in fermented juice by L. paracasei and L. rhamnosus but not L. helveticus (Table 2).…”

A Hydrolase from Lactobacillus sakei Moonlights as a Transaminase

“…The only meat-relevant Lactobacillus species for which a transaminase has been characterized until now is Lactobacillus paracasei. The enzyme showed temperature and pH optima at 43°C and pH 7.3, respectively (22). Comparatively, high temperatures seem to be essential for transamination in lactic acid bacteria.…”

“…Transamination activity assays of crude CFEs, purified enzyme (AcP), and lysozyme were performed as previously described (22), except that the concentrations varied. KP i (50 mM, pH 7.4), the substrate L-leucine (5 mM), ␣-ketoglutaric acid (5 mM), PLP (50 M), and 100 l protein solution in a total volume of 250 l were used.…”

“…Transaminases can be classified, among other methods, by specificity for branchedchain or aromatic amino acids. A transaminase from L. paracasei showed specificity only for BCAAs (22). Aromatic amino acid transaminases act on the three aromatic amino acids, and some of them, like transaminase D from E. coli (47) and a transaminase from L. lactis (46), also accept L-leucine and Lmethionine as substrates.…”