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PMID: 2646278 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S.

Analysis of prepro-alpha-lytic protease expression in Escherichia coli reveals that the pro region is required for activity.

Journal of bacteriology ·Vol. 171 ·No. 3 ·1989-03-00 ·Pages 1320-5

Silen JL, Frank D, Fujishige A, Bone R, Agard DA

Abstract

The alpha-lytic protease of Lysobacter enzymogenes was successfully expressed in Escherichia coli by fusing the promoter and signal sequence of the E. coli phoA gene to the proenzyme portion of the alpha-lytic protease gene. Following induction, active enzyme was found both within cells and in the extracellular medium, where it slowly accumulated to high levels. Use of a similar gene fusion to express the protease domain alone produced inactive enzyme, indicating that the large amino-terminal pro region is necessary for activity. The implications for protein folding are discussed. Furthermore, inactivation of the protease by mutation of the catalytic serine residue resulted in the production of a higher-molecular-weight form of the alpha-lytic protease, suggesting that the enzyme is self-processing in E. coli.

MeSH Terms
Cloning, Molecular Enzyme Precursors/genetics,isolation & purification,metabolism Escherichia coli/enzymology,genetics Genes Genes, Bacterial Genetic Vectors Gram-Negative Bacteria/enzymology,genetics Mutation Oligonucleotide Probes Plasmids Recombinant Proteins/isolation & purification,metabolism Serine Endopeptidases/genetics,isolation & purification,metabolism
Chemicals
Enzyme Precursors Oligonucleotide Probes Recombinant Proteins Serine Endopeptidases prepro-alpha-lytic protease myxobacter alpha-lytic proteinase
Authors & Affiliations
5 authors, click to expand affiliations / ORCID
Silen J L
Department of Biochemistry and Biophysics, University of California, San Francisco 94143-0448.
Frank D
Fujishige A
Bone R
Agard D A
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Article Info
Journal
Journal of bacteriology
Abbr.
J Bacteriol
ISSN
0021-9193
DOI
10.1128/jb.171.3.1320-1325.1989
Published
1989-03-00
Pages
1320-5
Language
English
Region
United States
NLM ID
2985120R
PMCID
PMC209748
Subset
IM
External Links
PubMed source DOI: 10.1128/jb.171.3.1320-1325.1989 PMC Full Text View journal details
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