[Thermoanaerobacter ethanolicus gene cluster containing the alpha- and beta-galactosidases genes melA and lacA, and properties of recombinant lacA].

The nucleotide sequence of a 4936 bp Thermoanaerobacter ethanolicus genomic DNA fragment containing the thermostable beta-galactosidase gene lacA and two incomplete open reading frames has been determined. The product of the first frame is highly homologous to alpha-galactosidases (melibiases), the product of the third frame is homologous to the alpha-D-mannosidases. The terminal area of the lacA, immediately following the stop-codon, harbors presumably a transcription termination site. Based on the location of the putative alpha-galactosidase gene melA and of the beta-galactosidase gene lacA on the T. ethanolicus chromosome, their combined transcription could be presumed. The calculated molecular mass of LacA is 86 kDa. LacA belongs to GH family 2 (GH2). Maximal activity of the purified recombinant enzyme was observed between pH values of 5.7 and 6.0 and temperatures of 75-80 degrees C. The highest activity, 480 units mg(-1), was found on lactose (Km 30 mM), the activities on pNPhGal and oNPhGal amounting to 330 and 420 units mg(-1), respectively. Immobilization on aldehyde silochrome increases the thermostability of the enzyme and keeps its high activity.