Development of Bile Salt-Resistant Leuconostoc citreum by Expression of Bile Salt Hydrolase Gene.
Probiotic bacteria must have not only tolerance against bile salt but also no genes for antibiotic resistance. Leuconostoc citreum is a dominant lactic acid bacterium in various fermented foods, but it is not regarded as a probiotic because it lacks bile salt resistance. Therefore, we aimed to construct a bile salt-resistant L. citreum strain by transforming it with a bile salt hydrolase gene (bsh). We obtained the 1,001 bp bsh gene from the chromosomal DNA of Lactobacillus plantarum and subcloned it into the pCB4170 vector under a constitutive P710 promoter. The resulting vector, pCB4170BSH was transformed into L. citreum CB2567 by electroporation, and bile saltresistant transformants were selected. Upon incubation with glycodeoxycholic acid sodium salt (GDCA), the L. citreum transformants grew and formed colonies, successfully transcribed the bsh gene, and expressed the BSH enzyme. The recombinant strain grew in up to 0.3% (w/v) GDCA, conditions unsuitable for the host strain. In in vitro digestion conditions of 10 mM bile salt, the transformant was over 67.6% viable, whereas only 0.8% of the host strain survived.
Brain Korea 21 Center for Bio-Resource Development, Division of Animal, Horticultural, and Food Sciences, Chungbuk National University, Cheongju 361-763, Republic of Korea.
Seung Kee
SK
Cho
Brain Korea 21 Center for Bio-Resource Development, Division of Animal, Horticultural, and Food Sciences, Chungbuk National University, Cheongju 361-763, Republic of Korea.
Soo Jin
SJ
Lee
Brain Korea 21 Center for Bio-Resource Development, Division of Animal, Horticultural, and Food Sciences, Chungbuk National University, Cheongju 361-763, Republic of Korea.
So-Yeon
SY
Shin
Brain Korea 21 Center for Bio-Resource Development, Division of Animal, Horticultural, and Food Sciences, Chungbuk National University, Cheongju 361-763, Republic of Korea.
Jin Seok
JS
Moon
Brain Korea 21 Center for Bio-Resource Development, Division of Animal, Horticultural, and Food Sciences, Chungbuk National University, Cheongju 361-763, Republic of Korea.
Ling
L
Li
Brain Korea 21 Center for Bio-Resource Development, Division of Animal, Horticultural, and Food Sciences, Chungbuk National University, Cheongju 361-763, Republic of Korea.
Wooha
W
Joo
Department of Animal Resources Science, Dankook University, Cheonan 330-714, Republic of Korea.
Dae-Kyung
DK
Kang
Brain Korea 21 Center for Bio-Resource Development, Division of Animal, Horticultural, and Food Sciences, Chungbuk National University, Cheongju 361-763, Republic of Korea.
Nam Soo
NS
Han
D000581:Amidohydrolases / Q000235:genetics / Q000378:metabolism*; D001647:Bile Acids and Salts / Q000378:metabolism / Q000633:toxicity*; D003001:Cloning, Molecular; D003470:Culture Media / Q000737:chemistry; D024881:Drug Resistance, Bacterial; D015870:Gene Expression; D048191:Lactiplantibacillus plantarum / Q000201:enzymology / Q000235:genetics; D007934:Leuconostoc / Q000187:drug effects* / Q000201:enzymology / Q000235:genetics / Q000254:growth & development*; D050296:Microbial Viability / Q000187:drug effects; D019936:Probiotics; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism*; D014170:Transformation, Genetic
D001647:Bile Acids and Salts; D003470:Culture Media; D011994:Recombinant Proteins; D000581:Amidohydrolases; C023154:choloylglycine hydrolase
Leuconostoc citreum; bile salt hydrolase; probiotics