RISTA AWALIA, - (2024) PEMURNIAN PROTEIN ALFA-AMILASE Saccharomycopsis fibuligera R64 MUTAN YANG DIEKSPRESIKAN DALAM INANG Pichia pastoris MENGGUNAKAN KROMATOGRAFI AFINITAS. Skripsi thesis, Sekolah Tinggi Farmasi Indonesia.
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Abstract
Pemurnian protein alfa-amilase sangat penting dalam industri untuk meningkatkan efisiensi dan kualitas produk. Oleh karena itu, penelitian ini memanfaatkan kromatografi afinitas dengan matriks Ni-Sepharose untuk memurnikan protein alfa-amilase. Pemurnian ini dilakukan dalam kondisi natif untuk mempertahankan struktur dan aktivitas biologis protein. Analisis elektroforesis SDS-PAGE digunakan untuk mengevaluasi keberhasilan pemurnian, sementara software ImageJ digunakan untuk menganalisis citra gel dari alfa-amilase, menentukan kemurnian, dan mengukur intensitas pita protein. Hasil analisis SDS-PAGE menunjukkan sebuah pita protein dominan pada ukuran ~48 kDa yang menandakan keberhasilan pemurnian. Aktivitas enzim diukur menggunakan metode Fuwa, dengan aktivitas tertinggi tercatat pada jam ke-144 induksi. Dengan kombinasi kromatografi afinitas dan analisis citra menggunakan Imagej menunjukkan bahwa pita dominan berada di ukuran sekitar ~48 kDa, dengan intensitas pita sebesar 208.27. Kemurnian protein dalam sampel mencapai 1,26% yang menunjukkan keberhasilan pemurnian enzim alfaamilase. Penelitian ini berhasil memurnikan alfa-amilase yang stabil dan aktif secara biologis. Hasil ini menunjukkan bahwa sistem ekspresi dan pemurnian yang digunakan efektif dalam menghasilkan alfa-amilase. ---- Alpha-amylase protein purification is very important in industry to improve efficiency and product quality. Therefore, this study utilized affinity chromatography with Ni-Sepharose matrix to purify alpha-amylase protein. This purification was carried out under native conditions to maintain the structure and biological activity of the protein. SDS-PAGE electrophoresis analysis was used to evaluate the success of the purification, while ImageJ software was used to analyze the gel image of alpha-amylase, determine the purity, and measure the intensity of the protein band. The results of the SDS-PAGE analysis showed a dominant protein band at a size of ~48 kDa indicating successful purification. Enzyme activity was measured using the Fuwa method, with the highest activity recorded at 144 hours of induction. The combination of affinity chromatography and image analysis using Imagej showed that the dominant band was around ~48 kDa, with a band intensity of 208.27. The purity of the protein in the sample reached 1.26% indicating the success of the purification of the alpha-amylase enzyme. This study succeeded in purifying stable and biologically active alpha-amylase. These results indicate that the expression and purification systems used are effective in producing alpha-amylase.
| Item Type: | Thesis (Skripsi) |
|---|---|
| Uncontrolled Keywords: | Alfa-amilase, Pichia pastoris, Pemurnian kromatografi afinitas, SDS-PAGE, Software ImageJ. ----- Alpha-amylase, Pichia pastoris, Affinity chromatography purification, SDS-PAGE, ImageJ software. |
| Subjects: | Q Science > Q Science (General) Q Science > QD Chemistry |
| Divisions: | Program Studi S1 Farmasi |
| Depositing User: | pustakawan - - |
| Date Deposited: | 13 Nov 2024 08:30 |
| Last Modified: | 13 Nov 2024 08:30 |
| URI: | http://repository.stfi.ac.id/id/eprint/1486 |
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