Amylolitic enzymes biosynthesis by submerged cultivation of Bacillus subtilis2+1

Bacillus subtilis je jedan od najvažnijih i najbolje proučavanih prokariota u prirodnim, biotehničkim i biomedicinskim znanostima jer se koristi kao modelni mikroorganizam u raznim znanstvenim istraživanjima. Proizvodnja amilaza submerznim uzgojem pomoću bakterijske vrste B. subtilis je počela još 1950. godine. Cilj ovog rada je pokazati kako se provodi biosinteza enzima α-amilaze te kojim se metodama ona pročišćava za daljnje korištenje. U ovom radu praćena je biosinteza i izolacija enzima α-amilaze iz bakterije Bacillus subtilis2+1 te je mjerena aktivnost enzima supernatantu i imobiliziranom enzimu Wohlgemuthovom metodom. Rezultati dobiveni u ovom radu pokazuju vrijeme maksimalne aktivnosti kulture Bacillus subtilis2+1, razliku u aktivnosti imobiliziranog i slobodnog enzima, razliku učinkovitosti metoda pročišćavanja enzima. Zaključeno je da proučavana bakterija proizvodi amilolitičke enzime, imobilizacijom se stabiliziraju enzimi i povećava njihova aktivnost, taloženje enzima etanolom je mnogo efikasnije od taloţenja enzima amonij-sulfatom. SDS-PAGE elektroforezom je ustanovljena molekulska masa enzima.Bacillus subtilis is one of the most important and best-studied prokaryotes in natural, biotechnical and biomedical science. It is the model miroorganism in scientific studies. Amylase production by submerged cultivation of bacteria, B. subtilis started in 1950. The aim of this paper is to show how the biosynthesis of an enzyme α-amylase is maintained, and the methods by which it is purified for further use. This paper deals with the biosynthesis of the enzyme, the isolation of an enzyme α-amylase from Bacillus subtilis2+1 and its activity which was measured by Wohgelmuth's method in immobilized form or in supernatant. The results obtained in this work presented the time of maximum activity of the culture Bacillus subtilis2+1, the distinction in the activity of immobilized and free enzymes and the efficiency of the methods of purification of an enzyme. By using them it is concluded that the studied bacteria produces amylolitic enzymes, that the immobilization is proven to stabilize enzymes, increasing their activity and that the precipitation of enzymes by ethanol is much more efficient than ammonium sulfate precipitation. Furthermore, the molecular mass of the enzyme as shown by SDS PAGE electrophoresis is shown