Solubilization and Functional Reconstitution of the Protein-translocation Enzymes of Escherichia Coli

Overview

Journal Proc Natl Acad Sci U S A

Specialty Science

Date 1990 Apr 1

PMID 2139227

Citations 9

Authors

A J Driessen

W Wickner

Affiliations

Soon will be listed here.

Abstract

The SecY protein and other membrane proteins of Escherichia coli were solubilized by mixed micelles of n-octyl beta-D-glucopyranoside, phospholipids, and glycerol. Proteoliposomes formed from this extract by detergent dialysis supported energy-dependent translocation and processing of pro-OmpA. Translocation required ATP, SecY, and SecA and was stimulated by a proton-motive force. These results provide an important assay for the isolation and identification of membrane components involved in protein translocation.

Citing Articles

Liposome technology for industrial purposes.

Wagner A, Vorauer-Uhl K J Drug Deliv. 2011; 2011:591325.

PMID: 21490754 PMC: 3065896. DOI: 10.1155/2011/591325.

A sporulation membrane protein tethers the pro-sigmaK processing enzyme to its inhibitor and dictates its subcellular localization.

Rudner D, Losick R Genes Dev. 2002; 16(8):1007-18.

PMID: 11959848 PMC: 152351. DOI: 10.1101/gad.977702.

Reconstitution of protein translocation from solubilized yeast membranes reveals topologically distinct roles for BiP and cytosolic Hsc70.

Brodsky J, Hamamoto S, Feldheim D, Schekman R J Cell Biol. 1993; 120(1):95-102.

PMID: 8416998 PMC: 2119491. DOI: 10.1083/jcb.120.1.95.

Interaction of wild-type signal sequences and their charged variants with model and natural membranes.

Rao N, Nagaraj R Biochem J. 1993; 293 ( Pt 1):43-9.

PMID: 8328971 PMC: 1134318. DOI: 10.1042/bj2930043.

Subunit dynamics in Escherichia coli preprotein translocase.

Joly J, Leonard M, Wickner W Proc Natl Acad Sci U S A. 1994; 91(11):4703-7.

PMID: 8197122 PMC: 43856. DOI: 10.1073/pnas.91.11.4703.

References

Rouser G, Fkeischer S, Yamamoto A . Two dimensional then layer chromatographic separation of polar lipids and determination of phospholipids by phosphorus analysis of spots. Lipids. 1970; 5(5):494-6. DOI: 10.1007/BF02531316. View

Lill R, Dowhan W, Wickner W . The ATPase activity of SecA is regulated by acidic phospholipids, SecY, and the leader and mature domains of precursor proteins. Cell. 1990; 60(2):271-80. DOI: 10.1016/0092-8674(90)90742-w. View

Chang C, Blobel G, Model P . Detection of prokaryotic signal peptidase in an Escherichia coli membrane fraction: endoproteolytic cleavage of nascent f1 pre-coat protein. Proc Natl Acad Sci U S A. 1978; 75(1):361-5. PMC: 411248. DOI: 10.1073/pnas.75.1.361. View

Newman M, Wilson T . Solubilization and reconstitution of the lactose transport system from Escherichia coli. J Biol Chem. 1980; 255(22):10583-6. View

Emr S, Silhavy T . Suppressor mutations that restore export of a protein with a defective signal sequence. Cell. 1981; 23(1):79-88. DOI: 10.1016/0092-8674(81)90272-5. View

Ito K . Purification of the precursor form of maltose-binding protein, a periplasmic protein of Escherichia coli. J Biol Chem. 1982; 257(17):9895-7. View

Oliver D, Beckwith J . Regulation of a membrane component required for protein secretion in Escherichia coli. Cell. 1982; 30(1):311-9. DOI: 10.1016/0092-8674(82)90037-x. View

Cerretti D, Dean D, Davis G, Bedwell D, Nomura M . The spc ribosomal protein operon of Escherichia coli: sequence and cotranscription of the ribosomal protein genes and a protein export gene. Nucleic Acids Res. 1983; 11(9):2599-616. PMC: 325911. DOI: 10.1093/nar/11.9.2599. View

Shiba K, Ito K, Yura T, Cerretti D . A defined mutation in the protein export gene within the spc ribosomal protein operon of Escherichia coli: isolation and characterization of a new temperature-sensitive secY mutant. EMBO J. 1984; 3(3):631-5. PMC: 557399. DOI: 10.1002/j.1460-2075.1984.tb01859.x. View

10.

Hjelmeland L, Chrambach A . Solubilization of functional membrane proteins. Methods Enzymol. 1984; 104:305-18. DOI: 10.1016/s0076-6879(84)04097-0. View

11.

Klausner R, van Renswoude J, Rivnay B . Reconstitution of membrane proteins. Methods Enzymol. 1984; 104:340-7. DOI: 10.1016/s0076-6879(84)04100-8. View

12.

Zwizinski C, Wickner W . The purification of M13 procoat, a membrane protein precursor. EMBO J. 1982; 1(5):573-8. PMC: 553089. DOI: 10.1002/j.1460-2075.1982.tb01210.x. View

13.

Rhoads D, Tai P, Davis B . Energy-requiring translocation of the OmpA protein and alkaline phosphatase of Escherichia coli into inner membrane vesicles. J Bacteriol. 1984; 159(1):63-70. PMC: 215593. DOI: 10.1128/jb.159.1.63-70.1984. View

14.

Muller M, Blobel G . Protein export in Escherichia coli requires a soluble activity. Proc Natl Acad Sci U S A. 1984; 81(24):7737-41. PMC: 392227. DOI: 10.1073/pnas.81.24.7737. View

15.

Wolfe P, Rice M, Wickner W . Effects of two sec genes on protein assembly into the plasma membrane of Escherichia coli. J Biol Chem. 1985; 260(3):1836-41. View

16.

Ito K . Identification of the secY (prlA) gene product involved in protein export in Escherichia coli. Mol Gen Genet. 1984; 197(2):204-8. DOI: 10.1007/BF00330964. View

17.

Driessen A, de Vrij W, Konings W . Incorporation of beef heart cytochrome c oxidase as a proton-motive force-generating mechanism in bacterial membrane vesicles. Proc Natl Acad Sci U S A. 1985; 82(22):7555-9. PMC: 391371. DOI: 10.1073/pnas.82.22.7555. View

18.

Geller B, Movva N, Wickner W . Both ATP and the electrochemical potential are required for optimal assembly of pro-OmpA into Escherichia coli inner membrane vesicles. Proc Natl Acad Sci U S A. 1986; 83(12):4219-22. PMC: 323703. DOI: 10.1073/pnas.83.12.4219. View

19.

Ambudkar S, Maloney P . Bacterial anion exchange. Use of osmolytes during solubilization and reconstitution of phosphate-linked antiport from Streptococcus lactis. J Biol Chem. 1986; 261(22):10079-86. View

20.

Bacallao R, Crooke E, Shiba K, Wickner W, Ito K . The secY protein can act post-translationally to promote bacterial protein export. J Biol Chem. 1986; 261(27):12907-10. View