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Modification of a Gene Cassette to Express Proglargine in Pichia pastoris: Elimination of Glu-Ala Repeats, Spacer and C-peptide Sequences Nasution, Uli Julia; Astuti, Rika Indri; Wahyudi, Aris Tri; Hardianto, Dudi; Martius, Efrida
Journal of Multidisciplinary Applied Natural Science Articles in Press
Publisher : Pandawa Institute

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.47352/jmans.2774-3047.368

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The growing global prevalence of diabetes mellitus has sharply increased the demand for insulin and its analogues. Pichia pastoris is a well-established system for recombinant pro-insulin and its analogues production. However, conventional gene constructs often include additional sequences, such as Glu-Ala repeats, spacer peptides, and c-peptides that complicate downstream processing and reduce efficiency. This study aimed to construct and express a proglargine (PG) gene cassette lacking the Glu-Ala repeats, spacer, and c-peptide in P. pastoris GS115 to obtain a uniform PG protein. The recombinant vector propagated in Escherichia coli TOP10F’, then expressed in P. pastorisGS115. Selected transformants were cultivated in YPG medium, then induced with 1% and 2% methanol daily in BMMY. The optimum methanol concentration further evaluated in ½ BSM induction medium. The result demonstrated that optimal PG expression was achieved with 2% methanol induction in BMMY, producing higher levels than those with ½ BSM. Among the transformants, PG.c2 produced the highest PG protein in BMMY medium induced with 2% methanol. Dot-blot analysis confirmed the secretion of PG, and LC-HRMS analysis demonstrated 100% amino acid sequence coverage of PG, confirming the integrity and completeness of the expressed protein. This study presented a newly modified PG gene cassette, inserted into pPICZαA vector, to express uniform secreted PG in P. pastoris GS115. By simplifying the precursor structure, a more homogeneous precursor product can be obtained, which is expected to simplify purification and also the downstream enzymatic process of PG into mature insulin glargine.
Extracellular Metabolites of Clove-Endophytic Bacteria, Niallia nealsonii DCL1, Exhibit Potential Cellular Antioxidant and Antiaging Activities Nadiyah Salsabilah; Muhammad Eka Prastya; Rika Indri Astuti
Biology, Medicine, & Natural Product Chemistry Vol 14, No 2 (2025)
Publisher : Sunan Kalijaga State Islamic University & Society for Indonesian Biodiversity

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.14421/biomedich.2025.142.747-755

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The clove plant, Syzygium aromaticum L., is a highly valued spice in Indonesia, recognized for its extensive applications in food preservation and medicinal uses. In a prior study, six endophytic bacteria were successfully isolated from clove leaves. One of these isolates, designated as DCL1, is a Gram-positive bacterium identified as Niallia nealsonii. This study aimed to determine the antioxidant and antiaging properties of the extracellular metabolite of DCL1. Extracellular metabolites were extracted with ethyl acetate and evaluated for antioxidant activity via the DPPH assay. Total phenolic and flavonoid compounds were also quantified with further metabolilte profiling using Liquid Chromatography Tandem-Mass Spectrometry (LC-MS/MS) analysis. Further study of antiaging activity was assayed using yeast model Schizosacchromyces pombe. The extract from N. nealsonii DCL1 cultures harvested at 28 and 40 hours yielded 0.01% and 0.009%, respectively. Our findings indicate that the 40-hour extract contains higher levels of phenolics (93.54 mg gallic acid equivalents/g extract) and flavonoids (12.09 mg quercetin equivalents/g extract) compared to the 28-hour extract. Additionally, the 40-hour extract exhibited greater antioxidant activity than the 28-hour extract, as demonstrated by DPPH (IC50 222 µg/mL) and ABTS (IC50 511.43 µg/mL) assays. Moreover, the 40-hour extract significantly prolonged the viability of S. pombe, suggesting its potential as an anti-aging agent. LC-MS/MS analysis identified several potential bioactive compounds, including gallic acid and N-carbamoyl-2,3-dihydroxybenzamide, which may contribute to its bioactivity. Ultimately, our study indicates that DCL1 40-h extract showed antiaging potential, which is promising as source and formulation of biopharmaceutical or cosmeceutical products. 
Unravelling Arbuscular Mycorrhizal Community in Kaolin Post-Mining and Tropical Heath Forest Ecosystems in Belitung Regency, Indonesia Muhammad Hilman Aulia; Nampiah Sukarno; Rika Indri Astuti; Adi Yulandi; Kartini Kramadibrata; Sri Listiyowati
HAYATI Journal of Biosciences Vol. 33 No. 4 (2026): July 2026
Publisher : Bogor Agricultural University, Indonesia

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.4308/hjb.33.4.851-863

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Arbuscular Mycorrhizal (AM) symbiosis plays a vital role in supporting plant growth in restored degraded land and natural forests. This study examined AM communities present within the root and the rhizosphere of kaolin post-mining land areas and tropical heath forests using both spore morphology and metagenomic approaches. Soil and plant root samples were collected from three distinct zones: the kaolin post-mining land, ecotone, and heath forest, and used to establish trap cultures with Pueraria javanica. Spore-based identification revealed nine AMF species belonging to five genera, with Acaulospora and Glomus being the most dominant. Metagenomic analysis of 18S rRNA sequences from colonized P. javanica roots revealed 33 AMF-associated Amplicon Sequence Variants (ASVs), dominated by Glomus and Rhizophagus. Diversity indices indicated medium diversity levels across all sites, with the ecotone showing the highest evenness and species richness. Differences in AM communities between morphological and molecular approaches highlight the importance of integrating both methods to understand their diversity comprehensively. This baseline information contributes to the potential use of AMF as bio-restoration agents in the kaolin post-mining ecosystems. To our knowledge, this is the first report of AM communities in the kaolin post-mining land and tropical heath forest.
Antibacterial Activity of Clove-derived Endophytic Bacteria of Niallia nealsonii DCL1 and Bacillus paranthracis DCL3 Nadya Nabila Qurani; Jepri Agung Priyanto; Rika Indri Astuti
Quagga: Jurnal Pendidikan dan Biologi Vol 18 No 1 (2026): QUAGGA : Jurnal Pendidikan dan Biologi
Publisher : Program Studi Pendidikan Biologi, Fakultas Keguruan dan Ilmu Pendidikan, Universitas Kuningan

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.25134/quagga.v18i1.470

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Pathogenic bacterial infection is still a major health problem that causes death. Endophytic bacteria in medicinal plants have the potential as antibacterial agents. Thus, endophytic bacteria in the medicinal plant, clove (Syzygium aromaticum), allow it to produce the same bioactive compounds as those contained in its host. This study aimed to analyze the antibacterial activity of five endophytic bacterial isolates from clove leaves. Antibacterial activity was evaluated using disc diffusion assay. Based on the results, isolate Niallia nealsonii DCL1 have strong antibacterial activity against Escherichia coli ATCC 8739, while Bacillus paranthracis DCL3 isolate has intermediate antibacterial activity against Pseudomonas aeruginosa ATCC 15442. In addition, extracellular metabolites of these two isolates of DCL1 and DCL3 which harvested at stationary phase showed antibacterial activity against and E. coli ATCC 8739 and P. aeruginosa ATCC 15442. These data indicate that the potential antibacterial compounds from DCL1 and DCL3 are considered extracellular secondary metabolites. Further analysis on the mode of antibacterial action should be carried out for further medical application.
Screening and characterization of sponge-associated bacteria from Seribu Island, Indonesia producing cellulase and laccase enzymes WENANG MAHARSIWI; RIKA INDRI ASTUTI; ANJA MERYANDINI; Aris Tri Wahyudi
Biodiversitas Journal of Biological Diversity Vol. 21 No. 3 (2020)
Publisher : Society for Indonesian Biodiversity

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.13057/biodiv/d210317

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Abstract. Maharsiwi W, Astuti RI, Meryandini A, Wahyudi AT. 2020. Screening and characterization of sponge-associated bacteria from Seribu Island, Indonesia producing cellulase and laccase enzymes. Biodiversitas 21: 975-981. Exploration of new enzymes from an extreme environment is important to improve industrial efficiency. This study aimed to get sponge-associated bacteria from Seribu Island with the capability to produce cellulase and laccase. These enzyme activities were indicated by the clear zones on CMC medium for cellulase and the reddish-brown zone on Guaiacol medium for laccase. About 100 of sponge-associated bacteria have been isolated from 5 marine sponges used SWC and NA modified media. As screened, one isolate (AGN89) could produce both enzymes and 11 isolates could produce cellulase. Quantitative analysis was performed using the DNS method and obtained the activities of 4 best cellulolytic isolates ranged from 0.04-0.06 UmL-1 and 0.70-1.18 UmL-1 in enzyme and specific activities, respectively. Gene-based determination for the isolate producing laccase resulted in a ±1100 bp amplicon fragment which identified as multicopper oxidase family protein. Based on the 16S-rRNA gene, AGN89 and these 4 cellulolytic isolates were identified as Pseudomonas luteola strain NBRC 103146, Bacillus aerius strain 24k, Pseudomonas aeruginosa strain DSM 50071, Mycobacterium maritypicum strain DSM 20578, and Brachybacterium conglomeratum strain J 1015. This result suggests that the sponge-associated bacteria from Seribu Island could become new enzymes producer for further applications in industry.
Evaluation of Indigenous Pichia kudriavzevii from cocoa fermentation for a probiotic candidate Rahayu Wulan; Rika Indri Astuti; Yaya Rukayadi; Anja Meryandini
Biodiversitas Journal of Biological Diversity Vol. 22 No. 3 (2021)
Publisher : Society for Indonesian Biodiversity

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.13057/biodiv/d220331

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Abstract. Wulan R, Astuti RI, Rukayadi Y, Meryandini A. 2021. Evaluation of Indigenous Pichia kudriavzevii from cocoa fermentation for a probiotic candidate. Biodiversitas 22: 1317-1325. Currently, probiotics are becoming a concern along with a healthy lifestyle awareness. Besides bacteria, yeast can be used as a probiotic candidate with specific functional properties. Research on yeast as a probiotic is still limited (except for Saccharomyces boulardii). Previous research has isolated yeasts from the cocoa fermentation process. This study aimed to evaluate the probiotics properties and antioxidant activity of yeast strains isolated from spontaneous cocoa fermentation in Sukabumi, Indonesia. Previous research has isolated 23 yeast strains from spontaneous cocoa (Theobroma cacao L.) fermentation. Of the 23 yeast strains isolated from cocoa fermentation, 22 strains showed negative hemolysis as an indicator of non-pathogenic properties. Ten yeast strains were able to grow at 37 °C and 41 °C, pH 3, 0.5% bile salts, had autoaggregation ability (63.99-95.33%), and co-aggregation with S. typhimurium ATCC14028 (>80%), as character requirement for probiotic candidates. The genetic identification of the ten yeast strains showed that they were 99% identical to Pichia kudriavzevii. Based on its antioxidant activity, the P. kudriavzevii 2P10 metabolites had the highest percentage of inhibition (68.51%) against DPPH free radicals and resistance to H2O2 oxidative stress up to 3 mM. It can be concluded that P. kudriavzevii 2P10 is a promising probiotic candidate for functional foods and health purposes.
Identification of lipase producing bacteria from palm oil sewage sludge processing plant at Malimping, Banten, Indonesia Ika Rahmatul Layly; ANJA MERYANDINI; IS HELIANTI; RIKA INDRI ASTUTI
Biodiversitas Journal of Biological Diversity Vol. 22 No. 10 (2021)
Publisher : Society for Indonesian Biodiversity

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.13057/biodiv/d221045

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Abstract. Layli IR, Meryandini A, Helianti I, Astuti RI. 2021. Identification of lipase-producing bacteria from palm oil sewage sludge processing plant at Malimping, Banten, Indonesia. Biodiversitas 22: 4512-4524. Lipase (triacylglycerol hydrolase E.C.3.1.1.3) is an enzyme that catalyzes the hydrolysis reaction of triacylglycerol into free fatty acids and glycerols. Lipase has broad application spectrum and unique catalytic mechanism, as one of important enzymes for industries, and almost all enzymes used for industries are originated from microbes. Identification of lipase-producing bacteria from palm oil processing sewage sludge was carried out to determine the potential of isolates and the application of lipase enzymes for industry. This study identified lipase-producing bacteria from the screening and isolation results from palm oil sewage sludge in Malimping-Banten. Two isolates were obtained from screening and isolation results. The morphological and observation and Gram staining method resulted in mlp-1 colony isolate was Gram-negative with basil cell shape, while mlp-2 colony isolate was Gram-positive with basil cell shape. Qualitative test on two isolates in tributyrin agar showed that two isolates produced clear zone or lipase enzyme. Molecular identification using 16SrRNA sequence showed that mlp-1 colony isolate was Chryseobacterium gleum, while mlp-2 colony was Bacillus velezensis. Literature study stated that Chryseobacterium gleum are pathogenic bacteria, resulting in uncontinued identification process, while Bacillus velezensis are non-pathogenic bacteria that have potential for industrial application. Identification of Bacillus velezensis was continued by performing biochemical test and inhibitory test. The continued identification results supported the molecular identification of lipase-producing enzyme for Bacillus velezensis.
Methanol pigment extracts derived from two marine actinomycetes exhibit antibacterial and antioxidant activities DOVA KELVIN MESRIAN; WIDYA ESTI PURWANINGTYAS; RIKA INDRI ASTUTI; AKHMAD ENDANG ZAINAL HASAN; Aris Tri Wahyudi
Biodiversitas Journal of Biological Diversity Vol. 22 No. 10 (2021)
Publisher : Society for Indonesian Biodiversity

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.13057/biodiv/d221037

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Abstract. Mesrian DK, Purwaningtyas WE, Astuti RI, Hasan AEZ, Wahyudi AT. 2021. Methanol pigment extracts derived from two marine actinomycetes exhibit antibacterial and antioxidant activities. Biodiversitas 22: 4440-4447. In recent years, natural pigments produced by marine microorganisms, especially actinomycetes, have become an important source of medically valuable drugs and other active compounds. Based on that crucial fact, this study aimed to determine the toxicity, antimicrobial, and antioxidant activities of methanolic pigment extract derived from marine actinomycetes HV11.P3 and SCA54.P2. Both extracts exhibited high toxicity activity based on Brine Shrimp Lethality Test (BSLT). The results showed the lethal concentration 50% (LC50) values of actinomycetes HV11.P3 and SCA54.P2 pigment crude extracts were 94.60 µg/mL and 131.22 mg/mL, respectively. The antimicrobial activity of those extracts was also tested against several microbes and showed the best results for inhibiting against Bacillus subtilis and Escherichia coli. Furthermore, we assessed the antioxidant activity using 2,2-diphenyl-1-picrylhydrazyl (DPPH) radicals and the results showed the inhibitory concentration 50% (IC50) values ??of actinomycetes HV11.P3 and SCA54.P2 pigment crude extracts were 231.08 µg/mL and 369.3 µg/mL, respectively. By using GC-MS analysis, we identified chemical compounds of actinomycete pigment crude extracts, revealing that the most dominant compounds were 1H-Purine-2,6-dione, 3,7-dihydro-1,3,7-trimethyl- (CAS) (44.65%) for HV11.P3 and 5-Methoxypyrrolidin-2-one (30.23%) for SCA54.P2. The 16S rRNA gene sequence analysis showed that actinomycete HV11.P3 and actinomycete SCA54.P2 had the closest similarity to Micromonospora chalcea strain 1464-217L and Micromonospora tulbaghiae strain TVU1, respectively. This study revealed that two pigment crude extracts derived from actinimycetes HV11.P3 and SVA54.P2 had antibacterial, antioxidant, and toxicity activities. The actinomycete SCA54.P2 pigment crude extract had the best activity as an antibacterial against E. coli ATCC 8739, while actnomycete HV11.P3 pigment crude extract had the best activity as an antioxidant.
An orange pigment from the marine bacterium Paracoccus haeundaensis SAB E11 as a prospective source of natural antioxidants HERMAWATY ABUBAKAR; RIKA INDRI ASTUTI; SRI LISTYOWATI; IRMANIDA BATUBARA; ARIS TRI WAHYUDI
Biodiversitas Journal of Biological Diversity Vol. 23 No. 9 (2022)
Publisher : Society for Indonesian Biodiversity

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.13057/biodiv/d230940

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Abstract. Abubakar H, Astuti RI, Listyowati S, Batubara I, Wahyudi AT. 2022. An orange pigment from the marine bacterium Paracoccus haeundaensis SAB E11 as a prospective source of natural antioxidants. Biodiversitas 23: 4730-4737. Bacterial pigment extracts are the source of many natural antioxidant substances. The present study aimed to assess the antioxidant activity of an orange pigment derived from a marine bacterium, identified as Paracoccus haeundaensis SAB E11, analyzed by 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2-azino-bis(ethylbenzothiazoline-6-sulfonic acid) (ABTS). Methanol-pigmented crude extract of P. haeundaensis SAB E11 exhibited the best DPPH scavenging activity with an IC50 value of 54.7 mg/mL, while the n-hexane crude pigment extract of the bacterium reduced ABTS optimally with an IC50 value of 24.46 mg/mL. The analysis of antioxidant compounds in P. haeundaensis SAB E11 was identified by combining liquid chromatography with tandem mass spectrometry (LCMS/MS). The five antioxidant compounds from methanol and n-hexane crude pigment extract of P. haeundaensis SAB E11 have been detected. Polyphenols were found to be the major compound, such as 3,5,3’,4’-tetrahydroxystilbene, scopoletin, and liquiritigenin. The other compounds were dihydroactinidiolide (terpenes) and ricinoleic acid (a fatty acid compound). The orange-colored extract of marine bacterium P. haeundaensis SAB E11 could be promising pharmacological capabilities as an antioxidant.
The potency of yellow pigment extract from the marine bacterium Pseudomonas oryzihabitans SAB E-3 as an antioxidant agent DELFIANI ANGGIAS PUTRI; RIKA INDRI ASTUTI; ARIS TRI WAHYUDI
Biodiversitas Journal of Biological Diversity Vol. 25 No. 6 (2024)
Publisher : Society for Indonesian Biodiversity

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.13057/biodiv/d250627

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Abstract. Putri DA, Astuti RI, Wahyudi AT. 2024. The potency of yellow pigment extract from the marine bacterium Pseudomonas oryzihabitans SAB E-3 as an antioxidant agent. Biodiversitas 25: 2565-2575. Metabolic processes in cells can produce byproducts in the form of Reactive Oxygen Species (ROS). Excessive amounts of ROS are one of the causes of oxidative stress. Antioxidants are known to act as free radical-scavenging compounds. Pigment-producing bacteria associated with sponges have been widely reported to have antioxidant compounds. In this study, the marine bacterium Pseudomonas oryzihabitans SAB E-3 produced intracellular yellow pigments with potential antioxidant activity. Therefore, this study aimed to assess the in vitro antioxidant and mitochondrial activity at the cellular level and identify chemical compounds. The intracellular pigments produced by P. oryzihabitans SAB E-3 were extracted using methanol, acetone, chloroform, and ethyl acetate. The methanol pigment extract contained the highest number of phenolic and flavonoid compounds, which may have contributed to its antioxidant activity. Furthermore, the active fraction obtained from the bioautographic TLC and harvested using preparative TLC had stronger antioxidant activity against DPPH radicals (75.71 µg/mL) than the crude pigment extract. However, the ABTS radical-reducing activity was lower (226.59 µg/mL) than the crude pigment extract. The maximum wavelength of the pigment extract was 453 nm, indicating the presence of carotenoids. In addition, the lowest concentration of the crude pigment extract (50 µg/mL) and selected active fractions (18.75 µg/mL) increased yeast Schizosaccharomyces pombe ARC039 tolerance to oxidative stress and induced mitochondrial activity. Active fraction one extract analyzed using LC-MS/MS contained piceatannol, resveratrol, isorhapontigenin, isoliquiritigenin, liquiritin, and 2-Omethylisohemigossylic acid lactone, which may contribute to its antioxidant activity. Therefore, the yellow pigment extract and the active fraction produced by P. oryzihabitans SAB E-3 are potential candidates for natural antioxidant sources.
Co-Authors A M Sabbah, Sajida ADI YULANDI Agung Priyanto, Jepri Ai Hertati, Ai AKHMAD ENDANG ZAINAL HASAN Akmaliyah, Rizna Alfaridza, Annisa Nourma ALFIYAN, ACHMAD Amani, Febriyanti Nur Andhika Rahesta Devi ANDREAS ADHI SATYA Anja Meryandini Antonius Suwanto Apon Zaenal Mustopa Aris Tjahjoleksono Aris Tri Wahyudi Armanisa, Khafifah Chairunnisa, Sheila Chihombori, Tatenda Calvin DAROJATUL ULYA DELFIANI ANGGIAS PUTRI Dina Aribah DINI NURDIANI DOVA KELVIN MESRIAN DOVA KELVIN MESRIAN Dudi Hardianto, Dudi EDI HUSEN Efrida Martius Ekawati, Nurlaili ESTI PUSPITASARI EVRICK YONANTIKO Fathiah, Muhammad Fadhil Fatimah Firyal Noviatanti Nabilah Hartono, Faisal Diniamal HERMAWATY ABUBAKAR Ika Rahmatul Layly Iman Rusmana IRA ERDIANDINI Irawan, Herman Irmanida Batubara IS HELIANTI Jendri Mamangkey Jepri Agung Priyanto Jun Nomura Kartini Kramadibrata Lenka, Audria Bayu Lisdar Idwan Sudirman Mahsunah, Anis Herliyati Manguntungi, Baso Martha, Ina MUHAMMAD EKA PRASTYA Muhammad Eka Prastya Muhammad Hilman Aulia Muhammad Zulfiqri, Aldy Mutiara, Ilma Nabila Tsoerayya Gustia Pudjas Nadila Fany Ashafani Nadiyah Salsabilah Nadya Nabila Qurani Nampiah Sukarno Nasution, Uli Julia NISA RACHMANIA MUBARIK Nurfadhilla Rahmadhani Nurhakiki Prastya, Muhammad Eka Puspitasari, Dian Japany Puspitasari, Esti RAHAYU WULAN Rahayu Wulan Rahayu Wulan Rakhman, Ira Meylan Nafarah Refli, Redoyan Rifqiyah Nur Umami, Rifqiyah Nur RIKA RAFFIUDIN Ririn Herni Putri Rustam, Yepy Hardi Sarah Asih Faulina Sarah Asih Faulina, Sarah Asih Satya, Andreas Adhi Sidiq, Muhammad Raffel SRI BUDIARTI SRI ESTUNINGSIH SRI LISTYOWATI Sri Budiarti Sri Budiarti Poerwanto Sri Estuningsih Sri Listiyowati Theresia Yolanda Avelina Trinugroho, Joko P Uci Cahlia WENANG MAHARSIWI WIDYA ESTI PURWANINGTYAS Wulan Tri Wahyuni Yaya Rukayadi YAYA RUKAYADI YAYA RUKAYADI YEPY HARDI RUSTAM Yohanes Bernadino Putera Saju Zulhaerati Zulhaerati