title: Efficient expression of acetylcholine-binding protein from Aplysia californica in Bac-to-Bac system.
abstract: The Bac-to-Bac baculovirus expression system can efficiently produce recombinant proteins, but the system may have to be optimized to achieve high-level expression for different candidate proteins. We reported here the efficient expression of acetylcholine-binding proteins from sea hares Aplysia californica (Ac-AChBP) and a convenient method to monitor protein expression level in this expression system. Three key factors affecting expression of Ac-AChBP were optimized for maximizing the yield, which included the cell density, volume of the infecting baculovirus inoculums, and the culturing time of postinfection. We have found it to reach a high yield of ∼5 mg/L, which needs 55 h incubation after infection at the cell density of 2 × 10(6) cells/mL with an inoculum volume ratio of 1 : 100. The optimized expression system in this study was also applied for expressing another protein Ls-AChBP from Lymnaea stagnalis successfully. Therefore, this established method is helpful to produce high yields of AChBP proteins for X-ray crystallographic structural and functional studies.
authors.affiliation: Key Laboratory of Tropical Biological Resources, Ministry of Education, Key Lab for Marine Drug of Haikou, Hainan University, Haikou, Hainan 570228, China.
authors.forename: Bo
authors.initials: B
authors.lastname: Lin
authors.affiliation: Key Laboratory of Tropical Biological Resources, Ministry of Education, Key Lab for Marine Drug of Haikou, Hainan University, Haikou, Hainan 570228, China.
authors.forename: Hailing
authors.identifier: 0000-0002-3403-1521
authors.initials: H
authors.lastname: Meng
authors.affiliation: Key Laboratory of Tropical Biological Resources, Ministry of Education, Key Lab for Marine Drug of Haikou, Hainan University, Haikou, Hainan 570228, China.
authors.forename: Hui
authors.initials: H
authors.lastname: Bing
authors.affiliation: Key Laboratory of Tropical Biological Resources, Ministry of Education, Key Lab for Marine Drug of Haikou, Hainan University, Haikou, Hainan 570228, China.
authors.forename: Dongting
authors.initials: D
authors.lastname: Zhangsun
authors.affiliation: Key Laboratory of Tropical Biological Resources, Ministry of Education, Key Lab for Marine Drug of Haikou, Hainan University, Haikou, Hainan 570228, China.
authors.forename: Sulan
authors.initials: S
authors.lastname: Luo
mesh_terms: D000109:Acetylcholine / Q000235:genetics* / Q000378:metabolism; D000818:Animals; D001048:Aplysia / Q000235:genetics* / Q000378:metabolism; D016367:Baculoviridae; D002352:Carrier Proteins / Q000096:biosynthesis*; D018360:Crystallography, X-Ray; D015967:Gene Expression Regulation, Viral
chemical_list: D002352:Carrier Proteins; D000109:Acetylcholine