Novel high-performance metagenome β-galactosidases for lactose hydrolysis in the dairy industry

Publication Type

Journal contribution (peer reviewed)

Authors

Erich, S.; Kuschel, B.; Schwarz, T.; Ewert, J.; Böhmer, N.; Niehaus, F.; Eck, J.; Lutz-Wahl, S.; Stressler, T.; Fischer, L.

Year of publication

2015

Published in

Journal of Biotechnology

Pubisher

Elsevier

DOI

10.1016/j.jbiotec.2015.06.411

Page (from - to)

27 - 37

The industrially utilised β-galactosidases from Kluyveromyces spp. and Aspergillus spp. feature undesirable kinetic properties in praxis, such as an unsatisfactory lactose affinity (K_M) and product inhibition (K_I) by galactose. In this study, a metagenome library of about 1.3 million clones was investigated with a three-step activity-based screening strategy in order to find new β-galactosidases with more favourable kinetic properties. Six novel metagenome β-galactosidases (M1–M6) were found with an improved lactose hydrolysis performance in original milk when directly compared to the commercial β-galactosidase from Kluyveromyces lactis (GODO-YNL2). The best metagenome candidate, called “M1”, was recombinantly produced in Escherichia coli BL21(DE3) in a bioreactor (volume 35 L), resulting in a total β-galactosidase M1 activity of about 1100 μkat_{oNPGal,37 °C} L⁻¹. Since milk is a sensitive and complex medium, it has to be processed at 5–10 °C in the dairy industry. Therefore, the β-galactosidase M1 was tested at 8 °C in milk and possessed a good stability (t_1/2 = 21.8 d), a desirably low apparent K_{M,lactose,8 °C} value of 3.8 ± 0.7 mM and a high apparent K_{I,galactose,8 °C} value of 196.6 ± 55.5 mM. A lactose hydrolysis process (milk, 40 nkat_lactose mL_milk,_8 °C⁻¹) was conducted at a scale of 0.5 L to compare the performance of M1 with the commercial β-galactosidase from K. lactis (GODO-YNL2). Lactose was completely (> 99.99 %) hydrolysed by M1 and to 99.6 % (w/v) by K. lactis β-galactosidase after 25 hours process time. Thus, M1 was able to achieve the limit of < 100 mg lactose per litre milk, which is recommended for dairy products labelled as “lactose-free”.