@article{lee_swaisgood_1998, title={Cloning and expression of a streptavidin-lipase fusion gene in Escherichia coli and characterization of the immobilized fusion protein}, volume={22}, ISSN={["1879-0909"]}, DOI={10.1016/S0141-0229(97)00174-9}, abstractNote={A gene for a streptavidin-lipase fusion protein was constructed by cloning a PCR-modified Pseudomonas fluorescens B52 lipase gene into a streptavidin chimeric protein expression vector, pStp4. The plasmids, pSTLP1 or pSTLP2, were used to transform Escherichia coli NM522 or DH5α. Plasmid pSTLP2, which contains a laclq gene upstream from the promoter, gave better expression of the bifunctional fusion protein when the cells were induced. Correct insertion of the fused gene was confirmed by DNA sequencing of the linker region and the lipase gene, by identification of a protein band of the correct molecular size (66 kDa) on SDS-PAGE, and by Western blot analysis using antistreptavidin antibody. Streptavidin-lipase fusion protein was purified and immobilized in a single step from recombinant cell lysates by bioselective adsorption on 6-(biotinamido)hexanamidopropyl-controlled pore glass. The hydrolytic activity of the resulting immobilized enzyme exhibited a pH optimum between 7 and 8 and a preference for short-chain fatty acids. Comparison of the hydrolytic activity of immobilized B52 lipase with that of commercial SAM-2 lipase from P. fluorescens suggests that about 30% of the activity was retained upon immobilization. Transesterification of tricaprylin with oleic acid in hexane by immobilized lipase indicated that 11% of the reaction products were derived from transesterification.}, number={4}, journal={ENZYME AND MICROBIAL TECHNOLOGY}, author={Lee, P and Swaisgood, HE}, year={1998}, month={Mar}, pages={246–254} }
 @article{lee_swaisgood_1997, title={Characterization of a chemically conjugated lipase bioreactor}, volume={45}, ISSN={["0021-8561"]}, DOI={10.1021/jf970167k}, abstractNote={Lipase from Candida cylindracea was immobilized on glass beads using the biospecific and high-affinity avidin−biotin interaction. Biotinylated lipase and glass beads were prepared by reactions of lipase and 3-aminopropyl glass beads with sulfosuccinimidyl-6-(biotinamido)hexanoate (NHS-LC-biotin). Avidin and biotinylated lipase were sequentially adsorbed to the biotinylated glass beads. Biotinylated lipase in solution retained about 63% of the hydrolytic specific activity of native lipase when an average 3 mol of biotin was incorporated/mol of lipase. Nonporous glass beads contained more biotin and protein (avidin and lipase) per unit of surface area, followed by 302 nm mean pore diameter controlled-pore glass beads (CPG-3000) and 198 nm mean pore diameter controlled-pore glass beads (CPG-2000). The hydrolytic specific activity of lipase immobilized on CPG-3000 and on nonporous beads was essentially the same as that for the biotinylated free enzyme, whereas that immobilized on CPG-2000 was about 50% less. ...}, number={8}, journal={JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY}, author={Lee, P and Swaisgood, HE}, year={1997}, month={Aug}, pages={3350–3356} }
 @article{lee_swaisgood_1997, title={Modification of milkfat physical properties by immobilized Pseudomonas fluorescens lipase}, volume={45}, ISSN={["0021-8561"]}, DOI={10.1021/jf970166s}, abstractNote={A bifunctional fusion protein was constructed by fusion of the streptavidin gene from Streptomyces avidinii to the lipase gene from Pseudomonas fluorescens, and the resulting streptavidin−lipase was expressed in Escherichia coli. Immobilized streptavidin−lipase was prepared by direct bioselective adsorption from crude cell lysates on biotinylated controlled-pore glass and used to catalyze interesterification of anhydrous butteroil. Changes in the triacylglycerol composition indicated that those with equivalent carbon numbers (ECN) ranging from 36 to 42 decreased, while those with ECN values from 48 to 50 increased following interesterification for 120 h in hexane at 42 °C. Both the melting temperatures and the solid fat content at various temperatures were lower as compared to those of the unmodified butteroil. Addition of unsaturated fatty acids, linoleic and linolenic, yielded modified butteroils with lower melting points and solid fat content, whereas addition of saturated fatty acids, palmitic and ste...}, number={8}, journal={JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY}, author={Lee, P and Swaisgood, HE}, year={1997}, month={Aug}, pages={3343–3349} }