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All Journal HAYATI Journal of Biosciences Jurnal Natural
Hartono, Faisal Diniamal
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Agriculture, Biological Sciences & Forestry Astronomy Biochemistry, Genetics & Molecular Biology Chemistry Earth & Planetary Sciences Energy Immunology & microbiology Neuroscience Physics

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Bioethanol Production from Non-Conventional Yeasts Wickerhamomyces anomalus (Pichia anomala) and Detection of ADH1 Gene Fathiah, Muhammad Fadhil; Hartono, Faisal Diniamal; Astuti, Rika Indri; Listiyowati, Sri; Meryandini, Anja
HAYATI Journal of Biosciences Vol. 31 No. 2 (2024): March 2024
Publisher : Bogor Agricultural University, Indonesia

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.4308/hjb.31.2.221-228

Abstract

Bioethanol is an organic compound resulted from the fermentation of sugar substrates by microorganisms which is used as alternative energy sources. During bioethanol fermentation yeast are exposed to various fermentation stresses, including temperature, osmotic, and oxidative stresess. Such conditions may decrease ethanol production. We previously isolated fermentation-stress tolerance yeast isolates from traditional Balinese beverages, identified as Wickerhamomyces anomalus BT2, BT5, and BT6. However no data available regarding the bioethanol production of those isolates. Our study indicates that these strains could utilize various sugar substrates (glucose, xylose, maltose, sucrose) in oxidative fermentative media. The highest value of substrate utilization efficiency following 48 hours fermentation was shown by BT6 on glucose (61.02%), BT 2 on xylose (55.44%) and maltose (60.90%). Measurement of ethanol production by Gas Chromatography showed that the strains were able to produce higher ethanol on the glucose substrate than other substrates. For instance, BT6 could produce the highest ethanol production (5.00 g/L) amongst strains tested by using glucose as substrate. Yet, the particular strains could only produce 0.30 g/L and 0.65 g/L by using xylose and maltose, respectively. For further genetic engineering purposes, we detected ADH1 gene from all three isolates, with high homology to the alcohol dehydrogenase from Saccharomyces cerevisiae, Geobacillus stearothermophilus and Pseudomonas aeruginosa. Further strain development can be carried out targeting the ADH1 gene, important for ethanol fermentation.
In silico characterization of adh1 gene encoding alcohol dehydrogenase 1 (ADH1) from non-conventional yeast, Wickerhamomyces and Pichia spp HARTONO, FAISAL DINIAMAL; MERYANDINI, ANJA; ASTUTI, RIKA INDRI
Jurnal Natural Volume 25 Number 3, October 2025
Publisher : Universitas Syiah Kuala

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.24815/jn.v25i3.45843

Abstract

Wickerhamomyces anomalus and Pichia kudriavzevii have high potential to produce bioethanol under high stress condition, due to their stress-tolerant properties. To elucidate and develop an efficient and sustainable bioethanol production, characterization of ethanol fermentation reactions is highly substantial. Ethanol fermentation employs key enzyme ADH1 encoded by ADH1 gene, important for conversion of acetaldehyde to ethanol. However, structural studies about alcohol dehydrogenase1 from these genera of yeasts are limited. This study aimed to detect the alcohol dehydrogenase 1 gene from Pichia spp. Using computational-bioinformatics approaches. The adh1 gene was amplified by PCR, visualized by electrophoresis, and analysed for sequence homology by BlastN and BlastP. The enzyme structure was constructed by SWISS-MODEL and I-TASSER with validation by Ramachandran plot, QMEAN4, and Local Quality Estimate. The Similarity and homology analysis of ADH1 genes and their corresponding protein sequence of yeast isolates showed that the ADH1gene was successfully detected. Multiple sequence alignment (MSA) and phylogenetic tree revealed that W. anomalus BT1-BT6 has close evolutionary relationship with ADH1 from Saccharomyces cerevisiae sequence while P. kudriavzevii IP4 showed different pattern. The ADH 1 enzyme model, generated using the SWISS-MODEL web server, demonstrated the best stereochemical quality, with a Ramachandran plot value of 100% for W. anomalus BT1 and 99.3% for P. kudriavzevii IP4. Superimposition of 3D-predicted model of ADH1 from W. anomalus BT1 and P. kudriavzevii 1P4 showed an exact match with amino acid in Zn2+ binding sites, confirming the ADH1 metaloenzyme properties. These findings provide structural insights about ADH1 genes and protein properties which can be used further for the development of efficient and high productivity of bioethanol productions through genetic and protein engineering.
Co-Authors Anja Meryandini Fathiah, Muhammad Fadhil Rika Indri Astuti Sri Listiyowati