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All Journal Makara Journal of Science Biosaintifika: Journal of Biology & Biology Education
Nataniel, Jocelyn
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Agriculture, Biological Sciences & Forestry

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Optimizing the Expression of Polyethylene Terephtalate Hydrolase-Encoding Synthetic Gene in Escherichia coli Arctic Express (DE3) Nataniel, Jocelyn; Ulfah, Maria; Achnafani, Dini; Nurhayati, Niknik; Sabbathini, Gabriela Christy; Wulandari, Sri Rezeki; Abinawanto, Abinawanto; Helianti, Is
Makara Journal of Science Vol. 28, No. 2
Publisher : UI Scholars Hub

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Abstract

The waste of polyethylene terephthalate (PET) plastic waste in Indonesia is a pressing concern due to its slow degradation and potential environmental damage. One promising solution is to utilize polyethylene terephthalate hydrolase from Ideonella sakaiensis (IsPETase), an enzyme that specifically degrades PET. However, inducing the expression of IsPETase synthetic gene in Escherichia coli BL21 (DE3) has been challenging because much of it remains insoluble. This study aimed to express IsPETase in E. coli Arctic Express (DE3) and optimize the conditions to enhance its production. First, pET22b(+)pelB-IsPETase was inserted into E. coli Arctic Express (DE3). The recombinant E. coli Arctic Express (DE3) was induced with isopropyl-β-D-1-thiogalactopyranoside (IPTG) and incubated at 10 °C. The fraction expressing soluble IsPETase was determined in different culture media, IPTG concentrations, induction times, and soni-cation durations. Parameters were optimized using a one-factor-at-a-time approach and then evaluated based on esterase specific activity and SDS-PAGE analysis. Results showed that IsPETase can be expressed in extracellular, periplasmic, and cytoplasmic soluble fractions. However, the extracellular fraction should be concentrated. Subsequent optimization focused only on the cytoplasmic fraction under optimal conditions, achieving a threefold increase in PETase specific activity compared with that under uninduced IPTG conditions. The reaction of PETase enzyme with PET and PCL was proven by weight loss, Scanning electron microscope (SEM), and Fourier transform infrared spectroscopy (FTIR). Although successful IsPETase expression and production optimization have been achieved, the specific activity remains low, prompting the need for ongoing expression optimization.
Valorization of Oil Palm Empty Fruit Bunch Waste Enhanced with Molasses for Erythritol Production Setiabang, Jeffersen Hosea; Fildy, Michael Ryan; Limawan, Kezia Zefanya; Widiyono, Advent Roan; Yonas, Nathanael Darrell; Nataniel, Jocelyn; Saputra, Indra Kurniawan; Kembaren, Riahna
Biosaintifika: Journal of Biology & Biology Education Vol. 17 No. 2 (2025): August 2025
Publisher : Universitas Negeri Semarang

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.15294/biosaintifika.v17i2.22551

Abstract

Indonesia, a leading palm oil producer, generates 1,250 tons of Oil Palm Empty Fruit Bunches (OPEFB) waste daily, typically disposed of through polluting combustion. This study explores repurposing OPEFB as a substrate for erythritol production, a popular sweetener with minimal impact on blood sugar. The research utilized Moniliella pollinis mutant SP5, a yeast capable of producing erythritol from various substrates. The process involved acid hydrolysis of OPEFB followed by fermentation. When fortified with 15% v/v molasses, OPEFB successfully served as an optimal carbon source for erythritol production, achieving a mass yield of 0.2878 g/g. Further improvement came through the use of ethyl methane sulfonate (EMS) mutagenesis, with the best mutant (mutant 7) reaching a yield mass of 0.3298 g/g and volumetric productivity of 0.0534 g/L/h. This research aims to advance Indonesia's self-sufficiency in erythritol production while providing a sustainable solution for OPEFB waste reduction, simultaneously addressing economic and environmental concerns.
Valorization of Oil Palm Empty Fruit Bunches (OPEFB) for Bioethanol Production in Indonesia Gozal, Celine; Halim, Amelia; Ridwan, Albert; Wong, Sidney Laura; Young, Kenneth Gabriel; Nataniel, Jocelyn; Kembaren, Riahna
Biosaintifika: Journal of Biology & Biology Education Vol. 16 No. 3 (2024): December 2024
Publisher : Universitas Negeri Semarang

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.15294/biosaintifika.v16i3.12099

Abstract

Indonesia, one of the world's largest producers of palm oil, generates large amounts of Oil Palm Empty Fruit Bunches (OPEFB). Its improper disposal can cause air pollution and soil acidification. OPEFB comprises lignocellulose, which can be hydrolyzed into monosaccharides to produce bioethanol. Bioethanol is a versatile product commonly produced using food crops as its raw material, but it can also be produced using lignocellulosic biomass to lessen the competition for food crops and land. Thus, this study aimed to produce bioethanol from OPEFB and enhance its production through adaptive evolution and supplementation with molasses. Adaptive evolution was carried out by exposing the yeast to ethanol concentrations of 3–12% (v/v). Native and adaptive strains are grown in OPEFB media supplemented with PEG and molasses. As a result, bioethanol was successfully produced, reaching around 10% (v/v) using a combination of OPEFB hydrolysate, PEG, and 20% molasses. Moreover, it was found that strains adapted toward 9% ethanol are more resistant to ethanol, as they can grow faster than the native strain. This shows that OPEFB can be effectively valorized for bioethanol production as a solution for OPEFB disposal and establishing a circular economy.
Co-Authors Abinawanto Abinawanto Achnafani, Dini Fildy, Michael Ryan Gozal, Celine Halim, Amelia Indra Kurniawan Saputra IS HELIANTI Kembaren, Riahna Limawan, Kezia Zefanya Maria Ulfah NIKNIK NURHAYATI Ridwan, Albert Sabbathini, Gabriela Christy Setiabang, Jeffersen Hosea Widiyono, Advent Roan Wong, Sidney Laura Wulandari, Sri Rezeki Yonas, Nathanael Darrell Young, Kenneth Gabriel