Exploring the acidotolerance of beta-galactosidase from Lactobacillus delbrueckii subsp. bulgaricus: an attractive enzyme for lactose bioconversion.
The LacZ gene encoding beta-galactosidase from Lactobacillus delbrueckii subsp. bulgaricus ATCC 11842 (L. bulgaricus) was cloned, sequenced and expressed in Escherichia coli, followed by purification and characterization of the protein. The recombinant enzyme was shown to be a homotetramer and could be distinguished from homologues by its relatively low and broad optimal temperature range, from 35 to 50 degrees C, coupled with an optimal pH of 5.0-5.5. Remarkably, the E491A mutant showed the same optimal temperature, but displayed an optimal pH at 6.5-7.0. Whilst these beta-galactosidases are inhibited by Cu(2+) they require only 1mM Mn(2+) and 1mM Co(2+) for optimal activity and thermostability. The wild-type enzyme was remarkably stable at acid pH values when compared to mutant E491A. Kinetic studies demonstrated that the E491A mutation affected catalysis rather than enzyme affinity. Furthermore, the wild-type protein efficiently cleaved lactose extracted from whey; however, in milk the E491A mutant showed the highest lactose bioconversion rate. Thus, these enzymes are interesting at the industrial level for hydrolysis of lactose extracted from whey or milk, and thus could contribute to overcoming the lactose intolerance problem generated by milk products.
Laboratoire d'Enzymes et de Métabolites des Procaryotes, Centre de Biotechnologie de Sfax, Route de Sidi Mansour Km 6 BP, 3038 Sfax, Tunisia.
Moez
M
Rhimi
Nushin
N
Aghajari
Bassem
B
Jaouadi
Michel
M
Juy
Samira
S
Boudebbouze
Emmanuelle
E
Maguin
Richard
R
Haser
Samir
S
Bejar
D001711:Biotransformation; D003001:Cloning, Molecular; D015870:Gene Expression; D005798:Genes, Bacterial; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis / Q000187:drug effects; D007763:Lac Operon; D048668:Lactobacillus delbrueckii / Q000201:enzymology* / Q000235:genetics; D007785:Lactose / Q000378:metabolism*; D017422:Sequence Analysis, DNA; D013696:Temperature; D001616:beta-Galactosidase / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*
D001616:beta-Galactosidase; D007785:Lactose