Identifikasi gen dan karakterisasi enzim α-amilase dari bacillus subtilis ifp1.1 = gene identification and characterization of α-amylase from bacillus subtilis ifp1.1

Fredrik, Mikhael (2019) Identifikasi gen dan karakterisasi enzim α-amilase dari bacillus subtilis ifp1.1 = gene identification and characterization of α-amylase from bacillus subtilis ifp1.1. Bachelor thesis, Universitas Pelita Harapan.

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Abstract

α-amylase enzyme is one of the most frequently used enzymes in the industry, due to its ability to hydrolyze starch at the liquefaction stage. The liquefaction stage is the most important stage in the process of producing sugar from the hydrolysis of starch. Testing the activity of α-amylase is important to determine the hydrolysis of α-amylase before it is used on a large scale. In this study, α-amylase extraction was produced by Bacillus subtilis IFP1.1. Furthermore, the protein concentration of the crude enzyme extract was determined by the Biuret method. Testing for α-amylase activity was carried out at optimum temperature, pH and substrate concentration using the reducing sugar (DNS) method. In addition, whole genome sequencing was carried out to determine the characterization of the α-amylase amino acid found in B. subtilis IFP1.1. Protein concentration in GYP medium was 1.39 unit/ mL, whereas in AYP medium it was 0.92 unit/ mL. The α-amylase produced from B. subtilis IFP1.1 works optimally at a temperature of 30 °C – 40 °C, pH 7, and a substrate concentration of 3%. Based on whole genome sequencing, the amino acid sequence α-amylase produced by B subtilis IFP1.1 has a percent matrix similarity of 98.79% with B. subtilis SUH4-2. In addition, there is no optimum temperature and pH difference between α-amylase produced from B. subtilis IFP1.1 and B. subtilis SUH4-2. The amino acid sequence of α-amylase shows that the catalyst side of B. subtilis IFP1.1 is in Aspartate 217, Glutamine 249, and Aspartate 310./ Enzim α-amilase merupakan salah satu enzim yang paling sering digunakan di industri, karena kemampuannya dalam menghidrolisis pati pada tahap likuifaksi. Tahap likuifaksi adalah tahap paling penting dalam proses produksi gula dari hidrolisis pati. Pengujian aktivitas α-amilase penting dilakukan untuk mengetahui kemampuan hidrolisis α-amilase sebelum digunakan dalam skala besar. Pada penelitian ini, dilakukan ekstraksi α-amilase yang dihasilkan oleh Bacillus subtilis IFP1.1. Selanjutnya, konsentrasi protein dari ekstrak enzim kasar tersebut ditentukan dengan metode Biuret. Pengujian aktivitas α-amilase dilakukan pada suhu, pH dan konsentrasi substrat optimum dengan metode gula pereduksi (DNS). Selain itu, whole genome sequencing dilakukan untuk mengetahui karakterisasi asam amino α-amilase yang terdapat pada Bacillus subtilis IFP1.1. Konsentrasi protein pada medium GYP sebesar 1,39 g/mL, sedangkan pada medium AYP sebesar 0,92 g/mL. Enzim α-amilase yang dihasilkan dari Bacillus subtilis IFP1.1 bekerja optimum pada suhu 30 °C – 40 °C, pH 7, dan konsentrasi substrat 3%. Berdasarkan whole genome sequencing, urutan asam amino α-amilase yang dihasilkan Bacillus subtilis IFP1.1 memiliki kemiripan persen matrix sebesar 98,79% dengan B. subtilis SUH4-2. Selain itu, tidak ada perbedaan suhu dan pH optimum antara α-amilase yang dihasilkan dari B. subtilis IFP1.1 dan B. subtilis SUH4-2. Urutan asam amino α-amilase menunjukkan bahwa sisi katalis dari B. subtilis IFP1.1 yaitu di Aspartat 217, Glutamin 249, dan Aspartat 310.

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