Previously, the author reported the purification of 5'-nucleotidase from bonito muscle by extraction with water, ammonium sulfate fractionation, DEAE-cellulose column chromatography and Sephadex G-200 gel filtration. In this paper, studies on some of the properties of the purified 5'-nucleotidase are reported.
This enzyme had a pH optimum of 5.5 in the presence of 0.01
M Mg
2+, with 5'-IMP as substrate. The rate of Pi liberation from 5'-IMP was Tinier for 80 minutes, the rate of hydrolysis decreasing slightly thereafter.
This enzyme showed highest relative activity with 5'-IMP as substrate, and also hydro-lyzed other 5'-nucleotides. On the other hand, it did not act on 3'(2')-nucleotides.
Of several metal ions Mg
2+ was most effective and Fe
2+ also activated this enzyme. Ca
2+, Mn
2+, Fe
3+ and Zn
2+ inhibited the 5'-nucleotidase activity to 56Y., 70%, 67% and 35 % of the control activity respectively, while Ba
2+ completely inhibited this enzyme.
The apparent Km value of 1.08 m
M and pKm of 2.97 were calculated for 5'-IMP. Using Sephadex G-200 gel filtration, the molecular weight of this enzyme was estimated to be 115, 000.
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