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Titolo:
Formation of soluble inclusion bodies by HPV E6 oncoprotein fused to maltose-binding protein
Autore:
Nomine, Y; Ristriani, T; Laurent, C; Lefevre, JF; Weiss, E; Trave, G;
Indirizzi:
Ecole Super Biotechnol Strasbourg, CNRS, UPR 9003, Lab RMN, F-67400 Illkirch Graffenstaden, France Ecole Super Biotechnol Strasbourg Illkirch Graffenstaden France F-67400 Ecole Super Biotechnol Strasbourg, UPRES 1329, Lab Immunotechnol, F-67400 Illkirch Graffenstaden, France Ecole Super Biotechnol Strasbourg Illkirch Graffenstaden France F-67400
Titolo Testata:
PROTEIN EXPRESSION AND PURIFICATION
fascicolo: 1, volume: 23, anno: 2001,
pagine: 22 - 32
SICI:
1046-5928(200110)23:1<22:FOSIBB>2.0.ZU;2-Z
Fonte:
ISI
Lingua:
ENG
Soggetto:
HUMAN-PAPILLOMAVIRUS TYPE-16; ESCHERICHIA-COLI; P53; ASSOCIATION; SOLUBILITY; FUSIONS; DOMAIN; PURIFICATION; DEGRADATION; THIOREDOXIN;
Keywords:
HPV oncoprotein E6; fusion protein; protein aggregation; protein overexpression; protein folding;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
34
Recensione:
Indirizzi per estratti:
Indirizzo: Trave, G Ecole Super Biotechnol Strasbourg, CNRS, UPR 9003, Lab RMN, F-67400 Illkirch Graffenstaden, France Ecole Super Biotechnol Strasbourg Illkirch Graffenstaden France F-67400
Citazione:
Y. Nomine et al., "Formation of soluble inclusion bodies by HPV E6 oncoprotein fused to maltose-binding protein", PROT EX PUR, 23(1), 2001, pp. 22-32

Abstract

Many polypeptides overexpressed in bacteria are produced misfolded and accumulate as solid structures called inclusion bodies. Inclusion-body-prone proteins have often been reported to escape precipitation when fused to maltose-binding protein (MEP). Here, we have examined the case of HPV 16 oncoprotein E6. The unfused sequence of E6 is overexpressed as inclusion bodies in bacteria. By contrast, fusions of E6 to the C-terminus of MBP are produced soluble. We have analyzed preparations of soluble MBP-E6 fusions by usingthree independent approaches: dynamic light scattering, lateral turbidimetry, and sandwich ELISA. All three methods showed that MBP-E6 preparations contain highly aggregated material. The behavior of these soluble aggregatesunder denaturating conditions suggests that they are formed by agglomeration of misfolded E6 moieties. However, precipitation is prevented by the presence of the folded and highly soluble MBP moieties, which maintain the aggregates in solution. Therefore, the fact that a protein or protein domain is produced soluble when fused to the C-terminus of a carrier protein does not guarantee that the protein of interest is properly folded and active. Wesuggest that aggregation of fusion proteins should be systematically assayed, especially when these fusions are to be used for binding measurements or activity tests. (C) 2001 Academic Press.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 03/07/20 alle ore 01:13:45