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J Bacteriol. May 1996; 178(10): 2986–2988.
PMCID: PMC178040

Site-specific proteolysis of the Escherichia coli SecA protein in vivo.


A seven-amino-acid cleavage site specific for tobacco etch virus (TEV) protease was introduced into SecA at two separate positions after amino acids 195 and 252. Chromosomal wild-type secA was replaced by these secA constructs. Simultaneous expression of TEV protease led to cleavage of both SecA derivatives. In the functional SecA dimer, proteolysis directly indicated surface exposure of the TEV protease cleavage sites. Cleavage of SecA near residue 195 generated an unstable proteolysis product and a secretion defect, suggesting that this approach could be used to inactivate essential proteins in vivo.

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Selected References

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  • Carrington JC, Dougherty WG. A viral cleavage site cassette: identification of amino acid sequences required for tobacco etch virus polyprotein processing. Proc Natl Acad Sci U S A. 1988 May;85(10):3391–3395. [PMC free article] [PubMed]
  • Dougherty WG, Cary SM, Parks TD. Molecular genetic analysis of a plant virus polyprotein cleavage site: a model. Virology. 1989 Aug;171(2):356–364. [PubMed]
  • Dougherty WG, Semler BL. Expression of virus-encoded proteinases: functional and structural similarities with cellular enzymes. Microbiol Rev. 1993 Dec;57(4):781–822. [PMC free article] [PubMed]
  • Driessen AJ. SecA, the peripheral subunit of the Escherichia coli precursor protein translocase, is functional as a dimer. Biochemistry. 1993 Dec 7;32(48):13190–13197. [PubMed]
  • Economou A, Wickner W. SecA promotes preprotein translocation by undergoing ATP-driven cycles of membrane insertion and deinsertion. Cell. 1994 Sep 9;78(5):835–843. [PubMed]
  • Guzman LM, Belin D, Carson MJ, Beckwith J. Tight regulation, modulation, and high-level expression by vectors containing the arabinose PBAD promoter. J Bacteriol. 1995 Jul;177(14):4121–4130. [PMC free article] [PubMed]
  • Ito K, Bassford PJ, Jr, Beckwith J. Protein localization in E. coli: is there a common step in the secretion of periplasmic and outer-membrane proteins? Cell. 1981 Jun;24(3):707–717. [PubMed]
  • Jarosik GP, Oliver DB. Isolation and analysis of dominant secA mutations in Escherichia coli. J Bacteriol. 1991 Jan;173(2):860–868. [PMC free article] [PubMed]
  • Kim YJ, Rajapandi T, Oliver D. SecA protein is exposed to the periplasmic surface of the E. coli inner membrane in its active state. Cell. 1994 Sep 9;78(5):845–853. [PubMed]
  • Kimura E, Akita M, Matsuyama S, Mizushima S. Determination of a region in SecA that interacts with presecretory proteins in Escherichia coli. J Biol Chem. 1991 Apr 5;266(10):6600–6606. [PubMed]
  • Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. [PubMed]
  • Matsuyama S, Kimura E, Mizushima S. Complementation of two overlapping fragments of SecA, a protein translocation ATPase of Escherichia coli, allows ATP binding to its amino-terminal region. J Biol Chem. 1990 May 25;265(15):8760–8765. [PubMed]
  • Mitchell C, Oliver D. Two distinct ATP-binding domains are needed to promote protein export by Escherichia coli SecA ATPase. Mol Microbiol. 1993 Nov;10(3):483–497. [PubMed]
  • Oliver DB. SecA protein: autoregulated ATPase catalysing preprotein insertion and translocation across the Escherichia coli inner membrane. Mol Microbiol. 1993 Jan;7(2):159–165. [PubMed]
  • Oliver DB, Beckwith J. E. coli mutant pleiotropically defective in the export of secreted proteins. Cell. 1981 Sep;25(3):765–772. [PubMed]
  • Parks TD, Leuther KK, Howard ED, Johnston SA, Dougherty WG. Release of proteins and peptides from fusion proteins using a recombinant plant virus proteinase. Anal Biochem. 1994 Feb 1;216(2):413–417. [PubMed]
  • Smith TA, Kohorn BD. Direct selection for sequences encoding proteases of known specificity. Proc Natl Acad Sci U S A. 1991 Jun 15;88(12):5159–5162. [PMC free article] [PubMed]
  • Weaver AJ, McDowall AW, Oliver DB, Deisenhofer J. Electron microscopy of thin-sectioned three-dimensional crystals of SecA protein from Escherichia coli: structure in projection at 40 A resolution. J Struct Biol. 1992 Sep-Oct;109(2):87–96. [PubMed]

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