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dc.contributor.author Kaderbhai, Mustak A.
dc.contributor.author Davey, Hazel M.
dc.contributor.author Kaderbhai, Naheed N.
dc.date.accessioned 2008-12-04T12:40:51Z
dc.date.available 2008-12-04T12:40:51Z
dc.date.issued 2004-06-09
dc.identifier.citation Kaderbhai , M A , Davey , H M & Kaderbhai , N N 2004 , ' A directed evolution strategy for optimized export of recombinant proteins reveals critical determinants for preprotein discharge ' Science , vol 13 , no. 9 , pp. 2458-2469 . , 10.1110/ps.04697304 en
dc.identifier.issn 1095-9203
dc.identifier.other PURE: 94187
dc.identifier.other dspace: 2160/1347
dc.identifier.uri http://hdl.handle.net/2160/1347
dc.identifier.uri http://www.proteinscience.org/cgi/reprint/13/9/2458 en
dc.description Mustak A. Kaderbhai, Hazel M. Davey and Naheed N. Kaderbhai (2004). A directed evolution strategy for optimized export of recombinant proteins reveals critical determinants for preprotein discharge. Protein Science, 13 (9) 2458-2469. RAE2008 en
dc.description.abstract A directed evolutionary approach is described that searches short, random peptide sequences for appendage at the secretory signal peptide–mature protein junction to seek ideal algorithms for both efficient and hyper export of recombinant proteins to the periplasm of Escherichia coli. The strategy employs simple, visual detection of positive clones using a PINK expression system that faithfully reports on export status of a mammalian hemoprotein in E. coli. With-in 'sequence spaces' ranging from 1 to 13 residues, a significant but highly variable secretory fitness was scored such that the rate of secretion reciprocally correlated with the membrane-associated precursor pool of the evolved exportable hemoproteins. Three clusters of hyper, median, and hypo exporters were isolated. These had corresponding net charges of –1, 0, and +1 within the evolved sequence space, which in turn clearly correlated with the prevailing magnitude and polarity of the membrane energization states. The findings suggest that both the nature of the charged residue and the proximal sequence in the early mature region are the crucial determinants of the protonophore-dependent electrophoretic discharge of the precursor across the inner membrane of E. coli. We conclude that the directed evolutionary approach will find ready application in engineering recombinant proteins for their efficient secretion via the sec export pathway in E. coli. en
dc.format.extent 12 en
dc.language.iso eng
dc.relation.ispartof Science en
dc.title A directed evolution strategy for optimized export of recombinant proteins reveals critical determinants for preprotein discharge en
dc.type Text en
dc.type.publicationtype Article (Journal) en
dc.identifier.doi http://dx.doi.org/10.1110/ps.04697304
dc.contributor.institution Institute of Biological, Environmental and Rural Sciences en
dc.description.status Peer reviewed en


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