• Journal of Life Science
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• v.29 no.4
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• pp.492-498
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• 2019

Previously, three kinds of lipases, lipAD1, lipAD2, and lipAD3, and lipase chaperone, lipBD, of Acinetobacter schindleri DYL129 isolated from soil sample were reported. In this report, three lipase and lipase chaperone were cloned into the pET32a(+) or pGEX-6P-1 vectors for protein expression in Escherichia coli, and named as pETLAD1, pETLAD2, pETLAD3 and pETLBD or pGEXLAD1, pGEXLAD 2, pGEXLAD3 and pGEXLBD, respectively. Protein expression rate was higher in pET system than in pGEX system. Although LipAD1 and LipAD2 were produced as inclusion bodies, their expression levels were high. So LipAD1 and LipAD2 could be solubilized in 8 M urea buffer and purified. LipAD3 and LipBD were overexpressed in soluble form and purified. Those proteins were purified by His-tag affinity chromatography connected in AKTA prime system. The activities of the purified lipases were demonstrated with 1% tributyrin agar plate. After purification, molecular mass was determined with sodium dodecyl sulfate-polyacrylamide gel electrophoresis. LipAD1 showed high activity toward ${\rho}$-nitrophenyl acetate and ${\rho}$-nitrophenyl butyrate, LipAD2 showed high activity toward ${\rho}$-nitrophenyl acetate and ${\rho}$-nitrophenyl myristate, and LipAD3 showed high activity toward ${\rho}$-nitrophenyl acetate, ${\rho}$-nitrophenyl butyrate, and ${\rho}$-nitrophenyl miristate, respectively. Three lipases, LipAD1, LipAD2, and LipAD3, showed optimal reaction at $50^{\circ}C$ using ${\rho}$-nitrophenyl butyrate, as substrate.

• The Korean Journal of Nuclear Medicine
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• v.30 no.1
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• pp.145-151
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• 1996

Pathways both mediated by and independent of transferrin(Tf) and the TfR have been described for the accumulation of iron. Although it is not clear whether the same systems take up iron and gallium, these pathways may suggest the contention that uptake of Ga-67 can, in fact, occur by both Tf-independent and Tf-dependent systems and may share with Fe-59 in part the same mechanism for uptake. The predominant system by which uptake of both radiometals occurs may be different in the degree of the transformation of tumor. Transformed(MMSV/3T3) and untransformed(BALB/3T3) cells were incubated with luM of Ga-67-citrate of Fe-59-chloride for 15 min. at $37^{\circ}C$ in either the presence or absence of Tf. After then, the monolayers were washed with HBSS or PBS, and the cells were solubilized in 1% SDS for gamma well counting and protein determinations. There were similarities, as well as differences, in the pattern of uptake of Fe-59 and Ga-67 presented both in ionic from and as bound to Tf. Both radiometals appeared gain to cells in either ionic or Tf-bound forms. Transformed cells appeared to accumulate more radiometal, either Ga-67 or Fe-59 in the presence of Tf than do the their untransforemd counterparts. Conversly the presentation of either radiometal in ionic form resulted in significantly greater accumulation of metal by the untransformed cells than those transformed. The efficiency for uptake of Ga-67 or Fe-59 in the absence of Tf was greater than for uptake of the Ga-Tf or Fe-Tf. However, the magnitude of difference in efficiency of uptake was greater for Fe-59(10-fold) than for Ga-67 (3-fold). Our results Supports the theory that both Tf-independent and Tf-dependent systems for the uptake of Ga-67 both systems operate oppositely between transformed cells and those untransformed, with uptake by the predominating in transformed cells by the Tf-mediated system and in untransformed cells by the Tf-independent. The uptake of Ga-67 by tumor may share with Fe-59 in part the same mechanism.