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. 1992 Mar 1;282(Pt 2):539–543. doi: 10.1042/bj2820539

On the catalytic mechanism of prokaryotic leader peptidase 1.

M T Black 1, J G Munn 1, A E Allsop 1
PMCID: PMC1130814  PMID: 1546969

Abstract

The catalytic mechanism of leader peptidase 1 (LP1) of the bacterium Escherichia coli has been investigated by a combination of site-directed mutagenesis, assays of enzyme activity in vivo utilizing a strain of E. coli which has a conditional defect in LP1 activity, and gene cloning. The biological activity of mutant forms of E. coli LP1 demonstrates that this enzyme belongs to a novel class of proteinases. The possibility that LP1 may be an aspartyl proteinase has been excluded on the basis of primary sequence comparison and mutagenesis. Assignment of LP1 to one of the other three recognized classes of proteinases (metalloproteinases, thiol proteinases and the classical serine proteinases) can also be excluded, as it is clearly demonstrated that none of the histidine or cysteine residues within LP1 are required for catalytic activity. The Pseudomonas fluorescens lep gene has been cloned and sequenced and the corresponding amino acid sequence compared with that of E. coli LP1. The E. coli LP1 and P. fluorescens LP1 primary sequences are 50% identical after insertion of gaps. The P. fluorescens LP1 has 39 fewer amino acids, a calculated molecular mass of 31903 Da and functions effectively in vivo in E. coli. None of the cysteine residues and only one of the histidine residues which are present in E. coli LP1 are conserved in sequence position in the P. fluorescens LP1 enzyme. The possibility that LP1 is a novel type of serine proteinase is discussed.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Baker R. K., Lively M. O. Purification and characterization of hen oviduct microsomal signal peptidase. Biochemistry. 1987 Dec 29;26(26):8561–8567. doi: 10.1021/bi00400a010. [DOI] [PubMed] [Google Scholar]
  2. Behrens M., Michaelis G., Pratje E. Mitochondrial inner membrane protease 1 of Saccharomyces cerevisiae shows sequence similarity to the Escherichia coli leader peptidase. Mol Gen Genet. 1991 Aug;228(1-2):167–176. doi: 10.1007/BF00282462. [DOI] [PubMed] [Google Scholar]
  3. Bilgin N., Lee J. I., Zhu H. Y., Dalbey R., von Heijne G. Mapping of catalytically important domains in Escherichia coli leader peptidase. EMBO J. 1990 Sep;9(9):2717–2722. doi: 10.1002/j.1460-2075.1990.tb07458.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Carter P., Wells J. A. Dissecting the catalytic triad of a serine protease. Nature. 1988 Apr 7;332(6164):564–568. doi: 10.1038/332564a0. [DOI] [PubMed] [Google Scholar]
  5. Dalbey R. E., Wickner W. Leader peptidase catalyzes the release of exported proteins from the outer surface of the Escherichia coli plasma membrane. J Biol Chem. 1985 Dec 15;260(29):15925–15931. [PubMed] [Google Scholar]
  6. Davies D. R. The structure and function of the aspartic proteinases. Annu Rev Biophys Biophys Chem. 1990;19:189–215. doi: 10.1146/annurev.bb.19.060190.001201. [DOI] [PubMed] [Google Scholar]
  7. Devereux J., Haeberli P., Smithies O. A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res. 1984 Jan 11;12(1 Pt 1):387–395. doi: 10.1093/nar/12.1part1.387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Greenburg G., Shelness G. S., Blobel G. A subunit of mammalian signal peptidase is homologous to yeast SEC11 protein. J Biol Chem. 1989 Sep 25;264(27):15762–15765. [PubMed] [Google Scholar]
  9. Higaki J. N., Evnin L. B., Craik C. S. Introduction of a cysteine protease active site into trypsin. Biochemistry. 1989 Nov 28;28(24):9256–9263. doi: 10.1021/bi00450a004. [DOI] [PubMed] [Google Scholar]
  10. Inada T., Court D. L., Ito K., Nakamura Y. Conditionally lethal amber mutations in the leader peptidase gene of Escherichia coli. J Bacteriol. 1989 Jan;171(1):585–587. doi: 10.1128/jb.171.1.585-587.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Lamotte-Brasseur J., Dive G., Dideberg O., Charlier P., Frère J. M., Ghuysen J. M. Mechanism of acyl transfer by the class A serine beta-lactamase of Streptomyces albus G. Biochem J. 1991 Oct 1;279(Pt 1):213–221. doi: 10.1042/bj2790213. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Miller M., Jaskólski M., Rao J. K., Leis J., Wlodawer A. Crystal structure of a retroviral protease proves relationship to aspartic protease family. Nature. 1989 Feb 9;337(6207):576–579. doi: 10.1038/337576a0. [DOI] [PubMed] [Google Scholar]
  13. Moore K. E., Miura S. A small hydrophobic domain anchors leader peptidase to the cytoplasmic membrane of Escherichia coli. J Biol Chem. 1987 Jun 25;262(18):8806–8813. [PubMed] [Google Scholar]
  14. Oefner C., D'Arcy A., Daly J. J., Gubernator K., Charnas R. L., Heinze I., Hubschwerlen C., Winkler F. K. Refined crystal structure of beta-lactamase from Citrobacter freundii indicates a mechanism for beta-lactam hydrolysis. Nature. 1990 Jan 18;343(6255):284–288. doi: 10.1038/343284a0. [DOI] [PubMed] [Google Scholar]
  15. Reid G. A., White S., Black M. T., Lederer F., Mathews F. S., Chapman S. K. Probing the active site of flavocytochrome b2 by site-directed mutagenesis. Eur J Biochem. 1988 Dec 15;178(2):329–333. doi: 10.1111/j.1432-1033.1988.tb14454.x. [DOI] [PubMed] [Google Scholar]
  16. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Wolfe P. B., Wickner W., Goodman J. M. Sequence of the leader peptidase gene of Escherichia coli and the orientation of leader peptidase in the bacterial envelope. J Biol Chem. 1983 Oct 10;258(19):12073–12080. [PubMed] [Google Scholar]
  18. YaDeau J. T., Klein C., Blobel G. Yeast signal peptidase contains a glycoprotein and the Sec11 gene product. Proc Natl Acad Sci U S A. 1991 Jan 15;88(2):517–521. doi: 10.1073/pnas.88.2.517. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. van Dijl J. M., van den Bergh R., Reversma T., Smith H., Bron S., Venema G. Molecular cloning of the Salmonella typhimurium lep gene in Escherichia coli. Mol Gen Genet. 1990 Sep;223(2):233–240. doi: 10.1007/BF00265059. [DOI] [PubMed] [Google Scholar]
  20. von Heijne G. Patterns of amino acids near signal-sequence cleavage sites. Eur J Biochem. 1983 Jun 1;133(1):17–21. doi: 10.1111/j.1432-1033.1983.tb07424.x. [DOI] [PubMed] [Google Scholar]

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