title: Creating an Efficient Methanol-Stable Biocatalyst by Protein and Immobilization Engineering Steps towards Efficient Biosynthesis of Biodiesel.
abstract: Two ternary sol-gel matrices, an octyltriethoxysilane-based aliphatic matrix and a phenyltriethoxysilane (PTEOS)-based aromatic matrix, were used to immobilize a methanol-stable variant of lipase from Geobacillus stearothermophilus T6 for the synthesis of biodiesel from waste oil. Superior thermal stability of the mutant versus the wildtype in methanol was confirmed by intrinsic protein fluorescence measurements. The influence of skim milk and soluble E. coli lysate proteins as bulking and stabilizing agents in conjunction with sol-gel entrapment were investigated. E. coli lysate proteins were better stabilizing agents of the purified lipase mutant than skim milk, as evidenced by reverse engineering of the aromatic-based system. This was also shown for commercial Candida antarctica lipase B (CaLB) and Thermomyces lanuginosus lipase (TLL). Uniform, dense, and nonaggregated particles imaged by scanning electron microscopy and a small particle size of 13 μm pertaining to the system comprising PTEOS and E. coli lysate proteins correlated well with high esterification activity. Combining protein and immobilization engineering resulted in a durable biocatalyst with efficient recycling ability and high biodiesel conversion rates.
authors: Department of Biotechnology and Food Engineering, Technion-Israel Institute of Technology, Haifa, 3200003, Israel.
authors: Shalev
authors: S
authors: Gihaz
authors: Department of Organic Chemistry and Technology, Budapest University of Technology and Economics, Műegyetem rkp. 3, H-1111, Budapest, Hungary.|Fermentia Microbiological Ltd., Berlini út 47-49, Budapest, H-1045, Hungary.|SynBiocat Ltd, Lövöház u 19/1, H-1023, Budapest, Hungary.
authors: Diána
authors: D
authors: Weiser
authors: Department of Biotechnology and Food Engineering, Technion-Israel Institute of Technology, Haifa, 3200003, Israel.
authors: Adi
authors: A
authors: Dror
authors: Fermentia Microbiological Ltd., Berlini út 47-49, Budapest, H-1045, Hungary.
authors: Péter
authors: P
authors: Sátorhelyi
authors: NanoTemper Technologies GmbH, Flößergasse 4, 81369, München, Germany.
authors: Moran
authors: M
authors: Jerabek-Willemsen
authors: Department of Organic Chemistry and Technology, Budapest University of Technology and Economics, Műegyetem rkp. 3, H-1111, Budapest, Hungary.|SynBiocat Ltd, Lövöház u 19/1, H-1023, Budapest, Hungary.|Biocatalysis and Biotransformation Research Group, Babes-Bolyai University of Cluj-Napoca, Arany János str. 11, RO-400028, Cluj-Napoca, Romania.
authors: László
authors: 0000-0002-8358-1378
authors: L
authors: Poppe
authors: Department of Biotechnology and Food Engineering, Technion-Israel Institute of Technology, Haifa, 3200003, Israel.
authors: Ayelet
authors: A
authors: Fishman
mesh_terms: D000818:Animals; D055162:Biocatalysis; D056804:Biofuels; D004795:Enzyme Stability / Q000187:drug effects; D004800:Enzymes, Immobilized / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry; D001411:Geobacillus stearothermophilus / Q000201:enzymology; D006868:Hydrolysis; D008049:Lipase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D000432:Methanol / Q000494:pharmacology*; D008892:Milk / Q000737:chemistry; D008958:Models, Molecular; D011487:Protein Conformation; D015202:Protein Engineering; D013696:Temperature
chemical_list: D056804:Biofuels; D004800:Enzymes, Immobilized; D029968:Escherichia coli Proteins; D008049:Lipase; D000432:Methanol
keywords: biodiesel; enzymes; immobilization; protein engineering; sol-gel processes