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酶的分子修飾backgrou-資料下載頁

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【正文】 es 2 mutagenic primers and 2 others。 ? Amplify a 5180。 fragment and a 3180。 fragment that overlap and each have the mutation。 ? Use these products in another reaction to produce the fulllength mutated DNA. PCR mutagenesis overlap extension SP6 primer forward mutagenic primer reverse mutagenic primer T7 primer X x x x Two separate 1st PCRs remove primers, denature and reanneal 2nd PCR “If a biochemist wishes to improve on a behaviour that is already optimized by natural selection, mutants are unlikely to yield enzymes with improved performances” S. Benner (1989) . 89,789806 . 肽鏈的有限水解與肽鏈的延伸生物體內：酶原激活主要是降低免疫原性 ?用木瓜蛋白酶水解亮氨酸氨肽酶， 2/3被除去，但活力不變 ?烯醇化酶在水解掉 150個氨基酸后，活力仍可保持肽鏈的延伸 Phusion iProof 通過物理方法改變酶的空間構象羧肽酶經高壓處理后，底物特異性發生改變，有利于合成反應纖維素酶高壓處理后，最適溫度降低，但活力提高補充材料 Esterification of phenylethanol with lauric acid CH 3HO? C C OC 11 H 23CH 3HOH? C + HOCOC 11 H 23EIs o o ct a n e40 oChr CH 3HOH? C + HOCOC11H23 CH 3HO? C C OC 11 H 23conversi243。n t (hours) 100 20 40 powder W/O O C O ( C H 2 ) 2 C O P E G NH P r tETwophases transgalactosilation ROH Gal OR Organic phase EE: ?galactosidase Glu Aqueous phase EGal?1,4Glu (lactose) conversi243。nt ( dias )2 4 6 8Gal ORROHER OHGal ORETwophases transgalactosilation ? Chemical modifications (in vitro engineering) ? One of the first way and a reemerged method for altering protein properties. ? Polyethylene glycol (PEG) modification of protein surface amino groups reduces immunoreactivity, prolongs clearance times, improve biostability, increase the solubility and activity of enzymes in anic solvents. ? (DeSantis, G. and Jones, . (1999) “Chemical modification of enzymes for enhanced functionality”, Current Opinion in Biotechnology, 10(4):324330) Some chemical modifications that may stabilize proteins Polyethylene glycol (PEG) modification of proteins Cyanuric chloride pnitrophenyl chloroformate

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