FOOD SCIENCE ›› 2020, Vol. 41 ›› Issue (2): 158-165.doi: 10.7506/spkx1002-6630-20181120-226

• Bioengineering • Previous Articles     Next Articles

Isolation, Identification and Enzymatic Characteristics of Kluyveromyces marxianus Producing β-D-Galactosidase

ZHENG Yi, ZHANG Jian, CAO Yongqiang, ZHAO Xiao, YU Zhijian, CHEN Chao, YANG Zhennai   

  1. (1. Beijing Advanced Innovation Center for Food Nutrition and Human Health, Beijing Technology and Business University, Beijing 100048, China; 2. Dongjun (Yucheng) Dairy Co. Ltd., Dezhou 253000, China)
  • Online:2020-01-25 Published:2020-01-19

Abstract: A strain, named ZX-5, with the ability to produce a high yield of β-D-galactosidase was isolated from traditional Tibetan kefir, and identified as Kluyveromyces marxianus by DNA internal transcribed spacer (ITS) sequence analysis. We found 2.0% galactose and 1.0% tryptone to be the optimal carbon and nitrogen sources, respectively. The optimal fermentation conditions were determined as follows: temperature 30 ℃, initial medium pH 6.5, medium volume 30%, shaking speed 100 r/min, inoculum size 2.0% and fermentation time 36 h. The activity of β-D-galactosidase produced under these conditions was 2.60 U/mL. Furthermore, the crude enzyme solution was purified by ammonium sulfate fractionation and DEAE-Sepharose fast flow ion-exchange chromatography to a specific activity of 157.35 U/mg. The purified β-D-galactosidase exhibited the maximum catalytic activity at 35 ℃ and pH 6.0, and had good stability at 20–40 ℃ and pH 5.0–7.0. Mn2+ could promote the enzyme activity. When the β-D-galactosidase produced by strain ZX-5 was used to decompose lactose and produce galacto-oligosaccharide under the conditions of 35 ℃, lactose concentration of 60 g/100 mL and enzyme concentration of 1.0 U/mL, the hydrolysis rate of lactose was 68.34% (after 50 h) and the yield of galacto-oligosaccharide was 34.70% (after 40 h), showing good potential application prospects.

Key words: Tibetan kefir, β-D-galactosidase, fermentation conditions, isolation and purification, enzyme characterization, galacto-oligosaccharide

CLC Number: