Thage B V, Rattray F P, Laustsen M W, Ardö Y, Barkholt V, Houlberg U
Department of Dairy and Food Science, Dairy Technology, The Royal Veterinary and Agricultural University, Rolighedsvej, Frederiksberg C, Denmark.
J Appl Microbiol. 2004;96(3):593-602. doi: 10.1111/j.1365-2672.2004.02163.x.
Purification and characterization of an aminotransferase (AT) specific for the degradation of branched-chain amino acids from Lactobacillus paracasei subsp. paracasei CHCC 2115.
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 alpha-ketoglutarate as the amino group acceptor; minor activity was shown for methionine. The enzyme had pH and temperature optima of 7.3 and 43 degrees C, respectively, and activity was detected at the pH and salt conditions found in cheese (pH 5.2, 4% NaCl). Hg2+ 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 carbonyl-binding 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.
The results suggest that Lact. paracasei subsp. paracasei CHCC 2115 may contribute to development of flavour in cheese.
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.
纯化并鉴定来自副干酪乳杆菌副干酪亚种CHCC 2115的、特异性降解支链氨基酸的转氨酶（AT）。
纯化方案包括阴离子交换色谱、亲和色谱和疏水相互作用色谱。该酶以单体形式存在，分子量为40 - 50 kDa。以α-酮戊二酸作为氨基受体时，该AT对异亮氨酸、亮氨酸和缬氨酸的转化速率相似；对蛋氨酸的活性较低。该酶的最适pH和温度分别为7.3和43℃，并且在奶酪中发现的pH和盐条件（pH 5.2，4% NaCl）下可检测到活性。Hg2+完全抑制该酶，在研究其他金属离子、巯基和羰基结合剂的作用时，其抑制模式与依赖磷酸吡哆醛的酶相似。该酶的N端序列为SVNIDWNNLGFDYMQLPYRYVAHXKDGVXD，在氨基酸水平上，与植物乳杆菌和乳酸乳球菌的支链氨基酸AT分别具有60%和53%的同一性。
结果表明副干酪乳杆菌副干酪亚种CHCC 2115可能有助于奶酪风味的形成。
这项工作的发现有助于了解与奶酪相关细菌进行的转氨作用，并有助于理解和控制氨基酸分解代谢以及香气化合物的产生。
