Extracellular Expression of Recombinant Kex2 Protease in Pichia pastoris BG10
Riyona Desvy PratiwiYuliawati YuliawatiFitriagustiani FitriagustianiAminah AminahHariyatun HariyatunNuruliawaty UtamiDini NurdianiAsrul Muhamad FuadM ManfrediA AntunesL JesusO DeM JulianoL JulianoW JudiceS DeY HuangY LongS LiT LinJ WuY ZhangY LinB GrangerM OutramP SolomonS WilliamsC OliveiraL DominguesD WaughS SreenivasS KrishnaiahN GovindappaY BasavarajuK KanojiaN MallikarjunJ NatarajanA ChatterjeeK SastryA Caballero-PrezJ Viader-SalvadA Herrera-EstalaJ Fuentes-GaribayM Guerrero-OlazarnN GovindappaM HanumanthappaK VenkatarangaiahK KanojiaK VenkatesanA ChatterjeeM KusumanchiN DaveP HazraS TiwariS YangY KuangH LiY LiuX HuiP LiZ JiangY ZhouY WangA XuG LukeM RyanN UtamiD NurdianiE Hariyatun; PutroW KusharyotoC KielkopfW BauerI UrbatschM TominagaK NozakiD UmenoJ IshiiA KondoT NaumannK SollenbergerG HaoN HigginsA VologodskiiS VermaZ QianS AdhyaM KarbalaeiS RezaeeH FarsianiR AwK PolizziQ ShenZ YuX.-T ZhouS.-J ZhangS.-P ZouN XiongY.-P XueZ.-Q LiuY.-G ZhengS ZhaoW GuanK MaY YanJ OuJ ZhangZ YuJ WuQ ZhangG LesageM TremblayJ GuimondG BoileauM Gmez-GaviriaN Lozoya-PrezM StaniszewskaB FrancoG Nio-VegaH Mora-MontesJ GaoL JiangJ LianZ Duman-zdamarB BinayL WohlschlagerF CsarmanM ZriliB SeibothR LudwigM NaseemG TajtiA GasparT SzantoJ BorregoG PanyiO KondrashovP KuzminS AkimovP AzaG MolpeceresF De SalasS CamareroP PuM Weissenborn
Abstract
Kex2 is a ~67 kD protease in yeast and fungi that specifically digest at -Lys-Arg- or -Arg-Arg-. Because of the same cleaving site, Kex2 is the potential to be applied in protein engineering, for instance, for protein purification. In the recent study, the Saccharomyces cerevisiae KEX2 gene was designed and constructed using pD902 and then integrated with the genome of Pichia pastoris BG10. Under AOX1 promoter control, Kex2 was expressed with methanol induction and found in extracellular fractions. The recombinant Kex2 was successfully confirmed with a western blot using a monoclonal anti-His-tag antibody and LC/MS/MS.
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