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PMID: 2527843 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S. Research Support, U.S. Gov't, P.H.S.

Genetic studies on the inability of beta-galactosidase to be translocated across the Escherichia coli cytoplasmic membrane.

Journal of bacteriology ·Vol. 171 ·No. 9 ·1989-09-00 ·Pages 4609-16

Lee C, Li P, Inouye H, Brickman ER, Beckwith J

Abstract

When a signal sequence is attached to beta-galactosidase, the normally cytoplasmic protein is unable to fully traverse the cytoplasmic membrane. We used a genetic approach to study those features of beta-galactosidase responsible for the block in translocation. By using both in vivo and in vitro techniques, fragments of beta-galactosidase were interposed between a signal sequence and alkaline phosphatase. The alkaline phosphatase acts as a sensor for any blocking effects of beta-galactosidase on export. From these studies, we show that multiple regions of beta-galactosidase contribute to its failure to be translocated. These results are most easily interpreted if the folding of beta-galactosidase or of domains of it is responsible for the block in export. In addition, in certain constructs, positively charged amino acids directly following the signal sequence interfered with export.

MeSH Terms
Alkaline Phosphatase/genetics Bacteriophage lambda/genetics Base Sequence Cell Membrane/metabolism Chromosome Deletion Escherichia coli/enzymology,genetics Galactosidases/genetics Genes Genes, Bacterial Genetic Vectors Genotype Molecular Sequence Data Plasmids Protein Processing, Post-Translational Protein Sorting Signals/genetics Recombinant Fusion Proteins/metabolism Signal Transduction Transduction, Genetic beta-Galactosidase/genetics,metabolism
Chemicals
Protein Sorting Signals Recombinant Fusion Proteins Alkaline Phosphatase Galactosidases beta-Galactosidase
Authors & Affiliations
5 authors, click to expand affiliations / ORCID
Lee C
Department of Microbiology and Molecular Genetics, Harvard Medical School, Boston, Massachusetts 02115.
Li P
Inouye H
Brickman E R
Beckwith J
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Article Info
Journal
Journal of bacteriology
Abbr.
J Bacteriol
ISSN
0021-9193
DOI
10.1128/jb.171.9.4609-4616.1989
Published
1989-09-00
Pages
4609-16
Language
English
Region
United States
NLM ID
2985120R
PMCID
PMC210258
Subset
IM
External Links
PubMed source DOI: 10.1128/jb.171.9.4609-4616.1989 PMC Full Text View journal details
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