purification and characterization of polyphenol oxidase from corn tassel(Stylus maydis)

In this work, purification of polifenol oxidase enzyme from stylus maydis was done and some kinetic characteristics were analysed. Polifenol oxidase extracted from stylus maydis was purified by the procedures of ammonium sulphate (NHࢩ(ࢧSOࢩprecipitation, dialysis, ultrafiltration and jel chromatography. One band for purified enzyme was obtained by SDSPAGE. Molecular weight of stylus maydis PPO enzyme was calculated as approximately 158 kDa. Optimum pH and temperature values of PPO enzyme were determined by using catechol and methyl catechol substrates. Polifenol oxidase activity was measured for 4-methyl catechol ve catechol at 420 nm as spectrophotometric. Optimum temperature was founded as 40°Cfor catechol and 30°C for 4-methyl catechol. Optimum pH was determined as 8 for catechol and 6 for 4-methyl catechol. Besides, substrat specifity and temperature inactivation of PPO were analysed. PPO enzyme's Km and Vmax values for catechol and 4-methyl catechol substrates at optimum pH and temperature were determined by using Linewear-Burk method. According to Vmax and Km values, the most suitable substrate was found to be catechol. Furthermore, it was observed that PPO activity decreased clearly in paralel with increasing inactivation time when catechol used. Inhibition effect of EDTA, SDS and sodium azide on PPO activity was analysed by using catechol substrate. In this work, sodium azide, EDTA and SDS were found to inhibit the enzyme activity as 26.6 %, 22.2 % and 12.2 %, respectively. Besides, the effects of carbohydrates such as sucrose, fructose, ribose and glucose on PPO activity were investigated. The enzyme was found to be activated 17 % by fructose and ribose, 16 % by glucose and 4 % by sucrose. Key words: Stylus maydis, polyphenol oxidase, substrate specifity, inhibition.