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β‐lactamase as a probe of membrane protein assembly and protein export
Author(s) -
BroomeSmith J. K.,
Tadayyon M.,
Zhang Y.
Publication year - 1990
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1111/j.1365-2958.1990.tb00540.x
Subject(s) - periplasmic space , biology , cytoplasm , escherichia coli , bacterial outer membrane , membrane protein , fusion protein , biochemistry , bacteria , gene , enzyme , microbiology and biotechnology , membrane , genetics , recombinant dna
Summary The enzyme TEM β‐lactamase constitutes a versatile gene‐fusion marker for studies on membrane proteins and protein export in bacteria. The mature form of this normally periplasmic enzyme displays readily detectable and distinctly different phenotypes when localized to the bacterial cytoplasm versus the periplasm, and thus provides a useful alternative to alkaline phosphatase for probing the topology of cytoplasmic membrane proteins. Celts producing translocated forms of β‐lactamase can be directly selected as ampicillin‐resistant colonies, and consequently a β‐lactamase fusion approach can be used for positive selection for export signals, and for rapid assessment of whether any protein expressed in Escherichia coli inserts into the bacterial cytoplasmic membrane. The level of ampicillin resistance conferred on a cell by an extracytoplasmic β‐lactamase derivative depends on its level of expression, and therefore a β‐lactamase fusion approach can be used to directly select for increased yields of any periplasmic or membrane‐bound gene products expressed in E. coli.