title: Molecular and enzymatic characterization of alkaline lipase from Bacillus amyloliquefaciens E1PA isolated from lipid-rich food waste.
abstract: Bacillus amyloliquefaciens E1PA is a lipase-producing strain that was originally isolated from lipid-rich food waste, and the production of its lipase was found to be induced by vegetable oils. The E1PA lipase was successfully expressed and secreted in a heterologous Escherichia coli host and was ultimately purified. The conserved pentapeptide motif Ala-His-Ser-Met-Gly was observed at positions 108-112. The purified recombinant lipase was stable over a pH range of 4.0-11.0 at 40 °C and exhibited maximal activity at pH 10. The recombinant E1PA lipase hydrolyzed a wide range of acyl esters (C4-C18). However, the highest activity (3.5 units mg(-1)) was observed when the p-nitrophenyl ester of myristate (C14) was used as a substrate. Compared to the lipases produced by Bacillus spp., the E1PA lipase displayed a structural molecular mass excluding the leader sequence (19.22 kDa) and a pI (9.82) that were similar to those reported for B. amyloliquefaciens lipases and lipase subfamily I.4 but that were quite distinct from those of lipase subfamily I.5 (approximately 43 kDa, pI 6). These results suggested that Bacillus lipases are closely related. Although the recombinant E1PA lipase digested only certain oils, the wild-type E1PA lipase degraded a variety of oils, including blended and re-used cooking oils. The recombinant and wild-type forms of the E1PA lipase were able to digest heterogeneous lipid-rich food waste at similar levels; this result suggests that this lipase can function even when it solely consists of its structural enzyme component. The enzyme exhibited lipid hydrolysis ability as either an intracellular domain of the recombinant protein or an extracellular domain secreted by the E1PA strain. However, the recombinant lipase showed higher activity than the wild-type E1PA lipase, indicating that the recombinant protein from E. coli possessed effective lipase activity. Thus, the inducible alkaline E1PA lipase exhibited the ability to act on a broad spectrum of substrates, and the effective form produced in the heterogeneous host can be further developed for several applications, such as biodiesel production and lipase production.
authors.affiliation: Department of Microbiology, Faculty of Science, Khon Kaen University, Khon Kaen 40002, Thailand.
authors.forename: Thanakorn
authors.initials: T
authors.lastname: Saengsanga
authors.affiliation: Department of Microbiology, Faculty of Science, Khon Kaen University, Khon Kaen 40002, Thailand; Center for Alternative Energy Research and Development, Khon Kaen University, Khon Kaen 40002, Thailand.
authors.forename: Wilailak
authors.initials: W
authors.lastname: Siripornadulsil
authors.affiliation: Department of Microbiology, Faculty of Science, Khon Kaen University, Khon Kaen 40002, Thailand; Center for Alternative Energy Research and Development, Khon Kaen University, Khon Kaen 40002, Thailand. Electronic address: surasak@kku.ac.th.
authors.forename: Surasak
authors.initials: S
authors.lastname: Siripornadulsil
mesh_terms: D000595:Amino Acid Sequence; D001407:Bacillus / Q000145:classification / Q000201:enzymology* / Q000235:genetics / Q000302:isolation & purification; D001426:Bacterial Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D004041:Dietary Fats / Q000378:metabolism* / Q000494:pharmacology; D004790:Enzyme Induction / Q000187:drug effects; D004926:Escherichia coli; D005516:Food Microbiology; D005798:Genes, Bacterial; D006868:Hydrolysis; D008049:Lipase / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D019216:Metals, Heavy / Q000494:pharmacology; D008958:Models, Molecular; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010802:Phylogeny; D010938:Plant Oils / Q000378:metabolism* / Q000494:pharmacology; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D062611:Solid Waste; D013045:Species Specificity; D013379:Substrate Specificity; D062065:Wastewater
chemical_list: D001426:Bacterial Proteins; D004041:Dietary Fats; D008049:Lipase; D019216:Metals, Heavy; D010938:Plant Oils; D011993:Recombinant Fusion Proteins; D062611:Solid Waste; D062065:Wastewater
keywords: Alkaline lipase; Bacillus amyloliquefaciens; Fatty acyl esters; Lipid-rich food wastes; Pentapeptide; Vegetable oils