Small single-chain variable fragments (scFv) are promising biomolecules to inhibit and neutralize toxins and to act as antivenoms. In this work, we aimed to produce a functional scFv-6009FV in the yeast Pichia pastoris, which inhibits the pure Cn2 neurotoxin and the whole venom of Centruroides noxius. We were able to achieve yields of up to 31.6 ± 2 mg/L in flasks. Furthermore, the protein showed a structure of 6.1 % α-helix, 49.1 % β-sheet, and 44.8 % of random coil by CD. Mass spectrometry confirmed the amino acid sequence and showed no glycosylation profile for this molecule. Purified scFv-6009FV allowed us to develop anti-scFvs in rabbits, which were then used in affinity columns to purify other scFvs. Determination of its half-maximal inhibitory concentration value (IC) was 40 % better than the scFvs produced by E. coli as a control. Finally, we found that scFv-6009FV was able to inhibit ex vivo the pure Cn2 toxin and the whole venom from C. noxius in murine rescue experiments. These results demonstrated that under the conditions assayed here, P. pastoris is suited to produce scFv-6009FV that, compared to scFvs produced by E. coli, maintains the characteristics of an antibody and neutralizes the Cn2 toxin more effectively.
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http://dx.doi.org/10.1016/j.ijbiomac.2024.133461 | DOI Listing |
Appl Biochem Biotechnol
January 2025
Unidad de Biotecnología Industrial, Centro de Investigación y Asistencia en Tecnología y Diseño del Estado de Jalisco A.C, 45019, Zapopan, Jal, Mexico.
Lipases from the basidiomycete fungus Ustilago maydis are promising but underexplored biocatalysts due to their high homology with Candida antarctica lipases. This study provides a comprehensive characterization of a recombinant CALB-like lipase from U. maydis, expressed in Pichia pastoris (rUMLB), and compares its properties with those of the well-studied recombinant lipase B from C.
View Article and Find Full Text PDFMicrob Biotechnol
January 2025
Izmir Biomedicine and Genome Center, Izmir, Turkey.
Low-cost and safe vaccines are needed to fill the vaccine inequity gap for future pandemics. Pichia pastoris is an ideal expression system for recombinant protein production due to its cost-effective and easy-to-scale-up process. Here, we developed a next-generation SARS-CoV2 Omicron BA.
View Article and Find Full Text PDFGlycobiology
January 2025
Department of Biochemistry, Dorothy Crowfoot Hodgkin Building, University of Oxford, South Parks Road, OX1 3QU, United Kingdom.
Abdala is a COVID-19 vaccine produced in Pichia pastoris and is based on the receptor-binding domain (RBD) of the SARS-CoV-2 spike. Abdala is currently approved for use in multiple countries with clinical trials confirming its safety and efficacy in preventing severe illness and death. Although P.
View Article and Find Full Text PDFInt J Biol Macromol
January 2025
State Key Laboratory of Biocatalysis and Enzyme Engineering, Hubei Key Laboratory of Industrial Biotechnology, School of Life Sciences, Hubei University, Wuhan 430062, PR China. Electronic address:
Expanding toolkits of EPA/DHA enrichment from natural sources is essential for better satisfying increasing demands for them. Lipase K80, from Proteus vulgaris K80, showed an application potential in EPA/DHA enrichment, whereas no desired heterologous expression in generally regarded as safe (GRAS) hosts restricted its relevant applications. In this study, expression of lipase K80 in a well-reputed GRAS host, Pichia pastoris, was achieved and further enhanced via combining disruption of its C-terminal KKL motif with co-expression of N-Acetyltransferase Mpr1, with a cumulative increment of nearly 200% in the secretion level and the volumetric activity.
View Article and Find Full Text PDFJ Agric Food Chem
January 2025
State Key Laboratory of Materials-Oriented Chemical Engineering, College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech University, Nanjing 211800, P.R. China.
Astaxanthin is a kind of carotenoid with a strong antioxidant ability, which has shown broad applications in the areas of healthcare, medicine, cosmetics, food additives, and aquaculture. With the increasing demand for natural products, the microbial production of astaxanthin has become a new hot spot. In this study, the astaxanthin synthesis pathway was first metabolically constructed in ()().
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