Mulay Y R, Deopurkar R L
Department of Microbiology, Tuljaram Chaturchand College, Baramati, Pune-413102, India.
Department of Microbiology, Savitribai Phule Pune University, Pune, 411007, India.
Appl Biochem Biotechnol. 2018 Feb;184(2):644-658. doi: 10.1007/s12010-017-2575-4. Epub 2017 Aug 23.
The amylase from Aureobasidium pullulans Cau 19 was purified by ammonium sulfate precipitation and Sephadex G-100 chromatography with a 9.25-fold increase in specific activity as compared to crude enzyme. Km and turn over values of the enzyme were 6.25 mg/mL and 5.0 × 10/min, respectively. Effect of different metal ions on the purified enzyme was investigated; 1 mM calcium (Ca) and cobalt (Co) enhanced enzyme activity by twofold; copper (Cu) had no effect on the activity of the enzyme. Mercury (Hg) 1 mM caused 90% inactivation whereas iron (Fe) and manganese (Mn) caused 10 to 16% inhibition. Amylase from A. pullulans Cau 19 was bioconjugated to gold nanoparticles synthesized using the biomass of A. pullulans Cau 19. Fourier transform infrared spectroscopy confirmed the conjugation of the enzyme to the gold nanoparticles. Though, only 20% of the added enzyme was adsorbed/conjugated on gold nanoparticles, 80% of the adsorbed activity could be estimated in the assay. The conjugated enzyme exhibited better tolerance to a broad pH range of 3.0-9.0 and higher temperatures compared with native enzyme.
对出芽短梗霉Cau 19产生的淀粉酶进行硫酸铵沉淀和Sephadex G - 100柱层析纯化，与粗酶相比，比活性提高了9.25倍。该酶的米氏常数（Km）和转换值分别为6.25 mg/mL和5.0×10/min。研究了不同金属离子对纯化后酶的影响；1 mM钙（Ca）和钴（Co）使酶活性提高了两倍；铜（Cu）对酶活性无影响。1 mM汞（Hg）导致90%的酶失活，而铁（Fe）和锰（Mn）导致10%至16%的抑制。将出芽短梗霉Cau 19产生的淀粉酶与利用出芽短梗霉Cau 19的生物质合成的金纳米颗粒进行生物偶联。傅里叶变换红外光谱证实了酶与金纳米颗粒的偶联。虽然，仅20%的添加酶吸附/偶联到金纳米颗粒上，但在测定中可估算出80%的吸附活性。与天然酶相比，偶联酶在3.0 - 9.0的宽pH范围和更高温度下表现出更好的耐受性。
