Purification and Characterization of the External Invertase Constitutively Produced by Rhodotorula glutinis K-24

Rhodotorula glutinis K-24에 의해 구성적으로 생산되는 세포외 Invertase의 정제 및 특성

Rhodoto& ghtbth~ K-24 was found to produce external invertase in addition to internal and cell wall bound invertase. External invertase was purified to an electrophoretically homogeneous state and partitally characterized and was compared with internal and cell wall bound invertase of which procedures for purification and characterization were reported previously. The enzyme was purified by ethanol precipitation, column chromatographies on DEAE-Sephadex A-50 and SP-Sephadex C-50, and gel filtration on Sephadex G-100. The molecular weight and subunit molecular weight of external invertasGwere estimated to be 220,000 and 100,000, respectively. The isoelectric point of the enzyme was about pH 6.0. The optimum pH and temperature for enzyme activity were pH 4.0 and $60^{\circ}C$, respectively. The enzyme remained stable at the wide range, from pH 3.0 to 11.0 and stable up to $40^{\circ}C$, but was inactivated at temperatures above that. $HgC_12, AgN0_3, MnS0_4$, SDS and p-CMB inhibited the enzyme activity. The $K_m$ value of the enzyme for sucrose was $1.0\times 10^{-2}$M. From these results, the three isozymes from Rh. glutinis K-24 seem to have the similar enzymatic properties, but to differ in molecular and subunit weights.

세포내 및 세포벽 뿐만 아니라, 세포외에도 invertase를 구성적으로 생산하는 효모 Rh.glutinis K-24로 부터 세포외 invertase를 disc 전기 영동상으로 단일한 상태로까지 정제하였다. 정제 효소의 효소화학적 성질을 밝힌 후 이미 보고한 바 있는 세포내 및 세포벽 invertase와 그 개락적 성질을 비교 검토하였다.