Casey Anne, Walsh Gary
Industrial Biochemistry Programme, Department of Chemical & Environmental Sciences, University of Limerick, Limerick city, Ireland.
Bioresour Technol. 2003 Jan;86(2):183-8. doi: 10.1016/s0960-8524(02)00145-1.
Extracellular phytase produced by Aspergillus niger ATCC 9142 was purified to homogeneity by employing an initial ultrafiltration step, followed by chromatography using ion exchange, gel filtration and chromatofocusing steps. The purified enzyme was an 84 kDa, monomeric protein. It possessed a temperature optimum of 65 degrees C, and a pH optimum of 5.0. Km and Vmax values of 100 microM and 7 nmol/s, respectively, were recorded and these values fall well within the range of those previously reported for microbial phytases. Substrate specificity studies indicated that, while the enzyme could hydrolyse a range of non-phytate-based phosphorylated substrates, its preferred substrate was phytate. Phytase activity was moderately stimulated in the presence of Mg2+, Mn2+, Cu2+, Cd2+, Hg2+, Zn2+ and F- ions. Activity was not significantly affected by Fe2- or Fe3- and was moderately inhibited by Ca2+. The enzyme displayed higher thermostability at 80 degrees C than did two commercial phytase products. Initial characterisation of the purified enzyme suggested that it could be a potential candidate for use as an animal feed supplement.
黑曲霉ATCC 9142产生的胞外植酸酶，首先通过超滤步骤进行初步纯化，然后依次经过离子交换色谱、凝胶过滤色谱和聚焦色谱步骤，最终纯化至同质。纯化后的酶是一种84 kDa的单体蛋白。其最适温度为65℃，最适pH值为5.0。记录的Km和Vmax值分别为100 microM和7 nmol/s，这些值完全落在先前报道的微生物植酸酶的范围内。底物特异性研究表明，虽然该酶能够水解一系列非植酸基的磷酸化底物，但其首选底物是植酸。在Mg2+、Mn2+、Cu2+、Cd2+、Hg2+、Zn2+和F-离子存在的情况下，植酸酶活性受到适度刺激。Fe2-或Fe3-对活性没有显著影响，而Ca2+则对其有适度抑制作用。该酶在80℃时表现出比两种市售植酸酶产品更高的热稳定性。对纯化酶的初步表征表明，它可能是用作动物饲料添加剂得潜在候选物。
