| Most thermostable enzymes are produced by thermophilic archaeaand they often have high thermostabilities and are resistant to organicsolvents or denaturants. Research on thermophilic archaea would notonly lend further evidence for biological evolution but also offerattractive industrial applications. Hyperthermophilic archaeonAeropyrum pernix K1 was isolated in 1993 from coastal solfotaricthermal vent in Kodakara-jima Island in Kyusyu, Japan..and was thefirst strictly aerobe archaea ever found and research on itsthermophilic enzymes would have vital meanings.Hyperthermophilic esterase APE1547 is from hyperthermophilicarchaea Aeropyrum pernix K1. It was cloned by PCR, and thenexpressed in the E. coli. BL-21 (DE) Codon Plus. The recombinantenzyme keeps its high thermostability. The We used the engineeringbacterium constructed by our lab to ferment and get the enzyme.Through the thermal denaturation, Hitrap Q Sepharose ion exchangechromatography and Sephacryl S-200 gel filtration chromatographywe got the superior thermophilic esterase APE of eletrophoreticpurity. Its proteic MW was determined by SDS-PAGE to be 63KDa.It is proved to be monomer by non-denatured electrophoresis andactive dying electrophoresis. The study for the zymologic charactersof the superior thermophilic esterase APE1547 shows that itsoptimum temperature is 90℃; its optimum PH is 8.0; its optimumsubstrate is p-nitrophenol octoate. In order to enlarge the applicationof the 12hyperthermophilic esterase APE1547 , increase the stabilityof using and preserving and find the stable mechanism of this enzymein the presence of extrinsic substract, in this paper ,we added somesugars ( lactose, glucose and sucrose ) and some polyols ( whichpossessing 2-6 hydroxys) to the enzyme to investigate the stability of... |
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