BACKGROUNDLipase is a versatile enzyme that serves as a biocatalyst in various industries. lk2 was successfully isolated from household waste compost through a metagenomic approach.MATERIALS AND METHODSlk2 from plasmid pPICZαA- lk2 was integrated into chromosomes of Pichia. pastoris GS115 using the electroporation method. Lk2 was expressed on Pichia. pastoris by methanol induction. The enzyme was purified through Ion Metal Affinity Chromatography Ni-NTA. The purified enzyme was characterized based on hydrolytic activity and in silico analysis.RESULTSLk2 was successfully expressed as an extracellular protein in Pichia pastoris. The cell-free supernatant exhibited hydrolysis activity to para-nitro phenyl laurate. The purified protein showed 15 times activity compared to cell-free supernatant and the size at around 35 kDa following gel electrophoresis. The enzyme showed optimum activity at 60^oC and pH 8. Lk2 preferred para nitro phenyl laurate as substrate. The enzyme's preference for medium-long carbon chains was corroborated by in silico analysis, which revealed favorable interactions between the enzyme and substrate, including affinity binding energy and optimal orientation of catalytic pocket to the substrate. Furthermore, the radius of gyration analysis of the Lk2 showed that the best structural compactness of Lk2 was at 60^oC. This is in line with the optimal temperature of Lk2 activity. In addition, docking analysis found important substrate binding residues, including Tyr30, Ser85, Leu121, Leu163, Leu166, Leu 233, and Leu254 beside Ser85, Asp231, and His253 as triad catalytic.CONCLUSIONLk2 belongs to a thermotolerant and alkaline lipase, prefers a medium-length carbon chain as substrate and is confirmed by in silico analysis. Several amino acid residues were probed to be important for substrate binding residues. The data give valuable information to develop the possibility of Lk2 as an industry's enzyme.

01 May 2025·3 Biotech

Silver/bacterial cellulose/Clitoria ternatea composite film for packaging application: synthesis, characterization and antibacterial properties

Article

Author: Andriani, Ade ; Noviana, Eka ; Indrianingsih, Anastasia Wheni ; Suwanda, Nadia Udanti ; Suryani, Ria ; Styaningrum, Putri ; Windarsih, Anjar

This study aimed to develop novel composite films made of Ag, bacterial cellulose (BC), and Clitoria ternatea flower extract for food packaging applications. The films were synthesized using varying concentrations of Ag ion (10^-3-10^-1 M) and flower extract (0-2x10⁴ µg/ml), followed by characterization using scanning electron microscopy-energy dispersive X-ray (SEM-EDX), X-ray diffraction (XRD), and Fourier-transform infrared (FTIR) spectroscopy. The antibacterial activity of the films was assessed against five bacteria, including Escherichia coli, Salmonella Typhimurium, Pseudomonas aeruginosa, Bacillus subtilis, and Staphylococcus aureus. The composite films appeared opaque and slightly dark. A porous network of microfibrils was observed under SEM, and higher Ag concentrations resulted in a rougher film surface. Higher Ag⁺ and extract concentrations resulted in higher percentages of Ag deposited on the surface, as confirmed by EDX (up to 34.7% at 2x10⁴ µg/ml extract and 10^-1 M Ag⁺). Ag/BC/C. ternatea extract combinations generally exhibit higher antibacterial activity than pure BC film. The highest inhibition of S. Typhimurium, P. aeruginosa, and B. subtilis was achieved by a composite film made with 10^-1 M Ag⁺ and 2x10⁴ µg/ml extract with inhibition values of 7.78 mm, 8.12 mm, and 8.25 mm, respectively. All tested composite films also had lower water vapor sorption capacity (2.31-6.71%, depending on the compositions) than BC (6.93%), enabling better protection of the preserved food from surrounding moisture. The Ag/BC/C. ternatea extract composite films are promising, sustainable packaging materials for preserving food quality.Supplementary InformationThe online version contains supplementary material available at 10.1007/s13205-025-04284-8.

01 May 2025·Current Microbiology

Complete Genome Sequence, Biological Activities, and Metabolomic Profiles of Mangrove-Derived Streptomyces sp. SM1P

Article

Author: Nofiani, Risa ; Mahmud, Taifo ; Juanssilfero, Ario Betha ; Siregar, Josephine Elizabeth ; Azmi, Wihda Aisarul ; Elfahmi ; Rizky ; Sukito, Agus ; Weisberg, Alexandra J ; Rizki, Andita Fitri Mutiara ; Syamsurizal ; Putri, Cantika Dylani ; Ardiningsih, Puji ; Rikayati, Rifa

The genome of Streptomyces sp. SM1P was sequenced to give a contig of 7,561,428 bp (71.7% G + C content) and 24 putative secondary metabolite biosynthetic gene clusters (BGCs). Eight of the BGCs showed high similarity (≥ 80%), three have moderate similarity (between 50 and 79%), and eleven showed low similarity (less than 50%) to known BGCs. The strain was cultivated in 7 different media prepared in 3 different solvents and the secondary metabolites were extracted with ethyl acetate to give 21 different extracts. All extracts showed antimicrobial and cytotoxic activities, with extracts from the ISP1/NaCl and ISP4/ASW cultures were the most active against bacteria and brine shrimp nauplii, respectively. The extract of the ISP2/NaCl culture, which gave a high number of spots on TLC, was also active against Plasmodium berghei. Untargeted metabolomic analysis of the ISP1/NaCl, ISP2/NaCl, and ISP4/ASW extracts using gas chromatography-mass spectrometry (GC-MS) and comparisons of the data against the National Institute of Standards and Technology 20 Mass Spectral Library showed variations between their secondary metabolites. Principal Component Analysis of the m/z values showed a correlation between ISP1/NaCl and ISP2/NaCl but not with ISP4/ASW. Metabolomic analysis of ISP2/NaCl extract using liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS) and comparisons of the data with those in the GNPS database and others indicated the presence of a lankamycin derivative (an antibiotic) and a cyclosporin derivative (an immunosuppressant). However, many other metabolites did not have a match. The results showed that Streptomyces sp. SM1P has potential as a producer of new secondary metabolites, particularly those with antimicrobial and antitumor activities.

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