sensitive　hydrogel　refolding　system：　preparation　of　poly（n-isopropyl　acrylamide）　and　its　application　in　lysozyme　refolding

《sensitive　hydrogel　refolding　system：　preparation　of　poly（n-isopropyl　acrylamide）　and　its　application　in　lysozyme　refolding》由会员上传分享，免费在线阅读，更多相关内容在教育资源-天天文库。

1、sensitive　Hydrogel　Refolding　System：　Preparation　of　Poly（N-isopropyl　acrylamide）　and　Its　Application　in　Lysozyme　RefoldingChineseJ.Chem.Eng.,12(4)556—560(2004)ATemperature—sensitiveHydrogelRefoldingPreparationofPoly(N—isopropylacrylamide)ItsApplicationinLysozymeRefoldingCUIZhifa

2、ng(崔志芳),DepartmentofChemicalandGuANYixin(关怡新)andBiochemicalEngineering,ZhejiangSystem:andYAOShanjing(y~善泾)University,Hangzhou310027,ChinaAbstractTemperature-sensitivehydrogel--poly(N-isopropylacrylamide)(PNIPA)waspreparedandappliedtoproteinrefolding.PNIPAgeldisksandgelparticlesw

3、eresynthesizedbythesolutionpolymerizationandinverseSUS.pensionpolymerizationrespectively.Theswellingkineticsofthegelswasalsostudied.WiththesepreparedPNIPAgels,themodelproteinlysozymewasrenatured.Within24h.PNIPAgeldisksimprovedtheyieldoflysozymeactivityby49.3%from3375.2U?mg一to503

4、8.8U?mg一.WiththeadditionoffasterresponsePNIPAgelbeads.thetotallysozymeactivityrecoverywasabout68.98%in3h,ascomparedwith42.O3%bvsimplebatchdilution.ThenovelrefoldingsystemwithPNIPAenablesemcientrefoldingespeciallyathighproteinconcentrations.Dis.cussionaboutthemechanismrevealedtha

5、twhenPNIPAgelswereaddedintotherefoldingbufrer.thehydrophobicinteractionsbetweendenaturedproteinsandpolymergelscouldpreventtheaggregationofrefoldingintermediates,thusenhancedtheproteinrenaturation.Keywordspoly(N-isopropylacrylamide)hydrogel,freeradicalpolymerization,proteinrefold

6、ing,lysozyme1INTRoDUCTIoNItiSknownthatheterologousproteinsproducedbyrecombinantDNAtechnologyareoffenpresentintheformofinactiveandinsolubleaggregatescalledinclusionbodies[11.Theseinsolubleaggregatesarethemisfoldedmolecules.Inordertoobtainproteinmoleculeswithbiologicalactivity.the

7、seinsolublepro—reinsmustbedissolvedbystrongdenaturantssuchasguanidinehydrochlorideorureaunderreducingcondition[.Withgraduallydecreasingconcentra.tionsofdenaturants,thedissolvedproteinmoleculesf_oldspontaneouslvintoitsnativeconformation.Usu.allytherefoldingofthedissolvedproteinin

8、toitsna.tiveconformationcanbeconsideredasaunimo