Uhland, Kerstin; Zander, Thomas; Ehrmann, Michael:
Synthetic competition between cytoplasmic folding and translocation of a soluble membrane protein domain
1995
In: Research in Microbiology, Jg. 146 (1995), Heft 2, S. 121 - 128
Artikel/Aufsatz in Zeitschrift / Fach: Biologie
Fakultät für Biologie
Titel:
Synthetic competition between cytoplasmic folding and translocation of a soluble membrane protein domain
Autor(in):
Uhland, Kerstin; Zander, Thomas; Ehrmann, Michael im Online-Personal- und -Vorlesungsverzeichnis LSF anzeigen
Erscheinungsjahr:
1995
Erschienen in:
Research in Microbiology, Jg. 146 (1995), Heft 2, S. 121 - 128
ISSN:
Signatur der UB:
Link URL:

Abstract:

In wild-type strains of Escherichia coli, alkaline phosphatase (AP), either when present as a soluble protein or when fused to a membrans protein, is only active after translocation to the periplasm. In thioredoxin reductase (trxB) mutants, however, cytoplasmically localized AP can form disulphide bonds and can reach an active conformation. Once it has folded in the cytoplasm, it can no longer be translocated. On the other hand, when AP is fused to periplasmic domains of a membrane protein, translocation can be more rapid than folding. Thus, expressing hybrids of AP and integral membrane proteins in a trxB mutant generates competition between folding of AP in the cytoplasm and its translocation to the periplasm. The cellular localization of AP can be monitored in phosphoserine phosphatase (serB) mutants causing auxotrophy for L-serine. Cytoplasmically but not periplasmically localized AP can compensate for the lack of SerB, leading to growth on indicator plates. As expected, when AP was fused to cytoplasmic domains of membrane proteins, serBmediated auxotrophy was abolished. Surprisingly, AP fusions to periplasmic domains exhibited a non-uniform response pattern. Fusions that translocate AP rapidly did not complement the SerB defect while those that export AP only slowly could do so. The usefulness of these strains for studying a variety of aspects related to membrane protein biogenesis is discussed.