대두의 발효를 통하여 생리활성을 가지고 있는 이소플라본 aglycone 함량을 높이기 위한 ${eta}$-glucosidase와 대두에 다량 함유되어 있는 stachyose, rafinose와 같은 난소화성 oligosaccharides를 분해하기 위해 ${alpha}$-galactosidase 효소 분비 미생물을 김치로부터 ${alpha}$-galactosidase와 ${eta}$-glucosidase를 생산하는 미생물을 탐색하였다. 탐색과정을 위해서 선별한 미생물을 16S rDNA sequencing 동정한 결과, Weissella cibaria 동정되어 Weissella cibaria K-M1-4로 명명하였다. Weissella cibaria K-Ml-4를 대두 액체배지에서 18시간동안 배양한 후, 생산한 효소는 배양액을 에탄을 침전, DEAE sepharose, sephacryl S-100HR column chromatography 통하여 ${alpha}$-galactosidase의 경우, 정제도 5.3배, 수율 3.5% 그리고 ${eta}$-glucosidase의 경우, 정제도 4.4배, 수율 2.9%로 정제되었다. ${alpha}$-Galactosidase 효소특성은 $60^{circ}C$에서 최대 활성을 나타내었으며, $80^{circ}C$에서 30분 처리시 43% 잔존활성을 보였다. pH 8.0에서 최대 활성을 나타내었으며, pH 5.0-9.0에서 안정하였다. 금속이온에 대한 영향에서 $Fe^{2+}$과 $Cu^{2+}$을 첨가하였을 때 효소 활성이 증가하였다. p-Nitrophenyl-${alpha}$-D-galacto-pyranoside (PNPG) 기질에 대한 Km은 0.98 mM이었고, Vmax는 $1.81{mu}$mole/min 이었다. ${eta}$-Glucosidase 효소 특성은 $50^{circ}C$에서 최대 활성을 나타내었으며, $80^{circ}C$에서 30분 처리시 46% 잔존활성을 보였다. pH 7.0에서 최대 활성을 나타내었으며, pH 5.0-9.0에서 안정하였다. 금속이온에 대한 영향에서 $Fe^{2+},;Co^{2+},;Cu^{2+}$을 첨가하였을 때 효소 활성이 증가하였다. p-Nitrophenyl-${eta}$-D-gluco-pyranoside (PNPG)에 대한 Km값은 1.24mM이었고, Vmax는 $6.81{mu}$mole/min 이었다.
A strain producing ${alpha}$-galactosidase and ${eta}$-glucosidase was isolated from Kimchi. The isolated strain was identified as Weissella cibaria by 16S rDNA analysis and designated as Weissella cibaria K-M1-4. The enzyme activity of ${alpha}$-galactosidase and ${eta}$-glucosidase reached the maximum in the soy medium at $37^{circ}C$ for 24 hr. The enzymes were purified by ethanol fractionation, DEAE sepharose fast flow, and sephacryl S-100HR column chromatography. ${alpha}$-Galactosidase specific activity was shown by 576 Units/mg protein and the yield was 3.5% of the total activity of crude extracts. ${eta}$-glucosidase specific activity was shown by 480 Units/mg protein and the yield was 2.9% of the total activity of crude extracts. The optimum temperature for ${alpha}$-galactosidase was $60^{circ}C$ and 43% of its original activity remained when it was treated at $80^{circ}C$ for 30 min. For ${alpha}$-galactosidase shows the optimum pH of 8.0 and is fairly stable between pH5.0 and pH9.0. The enzyme activity was increased in the presence of $Fe^{2+}$ and $Cu^{2+}$. The value of Km and Vmax for the enzyme were 0.98 mM and $1.81{mu}$mole/min, respectively. The ${eta}$-glucosidase has the optimum temperature of $50^{circ}C$ and 46% of its original activity remained when it was treated at $80^{circ}C$ for 30min. Its optimum pH of 7.0 and is fairly stable between pH5.0 and pH9.0. The enzyme activity was increased in the presence of $Fe^{2+},;Co^{2+}$ and $Cu^{2+}$. The value of Km and Vmax for the enzyme were 1.24 mM and $6.81{mu}$mole/min, respectively.
