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PMID: 8440256 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

An ATP transporter is required for protein translocation into the yeast endoplasmic reticulum.

The EMBO journal ·Vol. 12 ·No. 2 ·1993-02-00 ·Pages 659-66

Mayinger P, Meyer DI

Abstract

The transfer of precursor proteins through the membrane of the rough endoplasmic reticulum (ER) in yeast is strictly dependent on the presence of ATP. Since Kar2p (the yeast homologue of mammalian BiP) is required for translocation, and is an ATP binding protein, an ATP transport system must be coupled to the translocation machinery of the ER. We report here the characterization of a transport system for ATP in vesicles derived from yeast ER. ATP uptake into vesicles was found to be saturable in the micromolar range with a Km of 1 x 10(-5) M. ATP transport into ER vesicles was specifically inhibited by 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS), a stilbene derivative known to inhibit a number of other anion transporters, and by 3'-O-(4-benzoyl)benzoyl-ATP (Bz2-ATP). Inhibition of ATP uptake into yeast microsomes by DIDS and Bz2-ATP blocked protein translocation in vitro measured co- as well as post-translationally. The inhibitory effect of DIDS on translocation was prevented by coincubation with ATP. Moreover, selective membrane permeabilization, allowing ATP access to the lumen, restored translocation activity to DIDS-treated membranes. These results demonstrate that translocation requires a DIDS and Bz2-ATP-sensitive component whose function is to transport ATP to the lumen of the ER. These findings are consistent with current models of protein translocation in yeast which stipulate the participation of Kar2p in the translocation process.

MeSH Terms
4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid 4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid/analogs & derivatives,pharmacology Adenosine Triphosphate/analogs & derivatives,chemistry,metabolism Biological Transport/drug effects Carrier Proteins/metabolism Endoplasmic Reticulum/metabolism Fungal Proteins/metabolism HSP70 Heat-Shock Proteins Heat-Shock Proteins/metabolism Kinetics Microsomes/metabolism Photochemistry Protein Precursors/metabolism Saccharomyces cerevisiae/metabolism
Chemicals
Carrier Proteins Fungal Proteins HSP70 Heat-Shock Proteins Heat-Shock Proteins KAR2 protein, yeast Protein Precursors 4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid Adenosine Triphosphate 4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Mayinger P
Department of Biological Chemistry, UCLA School of Medicine.
Meyer D I
References (42)
42 references, click to expand
  1. Yeast Sec proteins interact with polypeptides traversing the endoplasmic reticulum membrane.
    Cell. 1992 Apr 17;69(2):343-52 PMID: 1568249
  2. Sec61p and BiP directly facilitate polypeptide translocation into the ER.
    Cell. 1992 Apr 17;69(2):353-65 PMID: 1568250
  3. Protein import into chloroplasts requires a chloroplast ATPase.
    Proc Natl Acad Sci U S A. 1987 May;84(10):3288-92 PMID: 2953027
  4. Rate-limiting steps in metabolic pathways.
    J Biol Chem. 1979 Mar 25;254(6):1875-8 PMID: 422559
  5. Secretion in yeast: preprotein binding to a membrane receptor and ATP-dependent translocation are sequential and separable events in vitro.
    J Cell Biol. 1989 Jun;108(6):2101-6 PMID: 2544601
  6. Mechanisms of membrane protein insertion into liposomes during reconstitution procedures involving the use of detergents. 1. Solubilization of large unilamellar liposomes (prepared by reverse-phase evaporation) by triton X-100, octyl glucoside, and sodium cholate.
    Biochemistry. 1988 Apr 19;27(8):2668-77 PMID: 2840945
  7. A protein-conducting channel in the endoplasmic reticulum.
    Cell. 1991 May 3;65(3):371-80 PMID: 1902142
  8. Protein translocation into the endoplasmic reticulum: a light at the end of the tunnel.
    Trends Cell Biol. 1991 Dec;1(6):154-9 PMID: 14731858
  9. Rapid and efficient site-specific mutagenesis without phenotypic selection.
    Methods Enzymol. 1987;154:367-82 PMID: 3323813
  10. A microsomal protein is involved in ATP-dependent transport of presecretory proteins into mammalian microsomes.
    EMBO J. 1991 Oct;10(10):2795-803 PMID: 1833183
  11. Effect of atractylosides, palmitoyl coenzyme A, and anion transport inhibitors on translocation of nucleotide sugars and nucleotide sulfate into Golgi vesicles.
    J Biol Chem. 1984 Apr 10;259(7):4263-6 PMID: 6707004
  12. Secretion in yeast: translocation and glycosylation of prepro-alpha-factor in vitro can occur via an ATP-dependent post-translational mechanism.
    EMBO J. 1986 May;5(5):1031-6 PMID: 15957217
  13. Signal recognition particle (SRP) stabilizes the translocation-competent conformation of pre-secretory proteins.
    EMBO J. 1988 Nov;7(11):3553-7 PMID: 2850167
  14. Synthesis and use of azido photoaffinity analogs of adenine and guanine nucleotides.
    Methods Enzymol. 1979;56:642-53 PMID: 459886
  15. Requirement for hsp70 in the mitochondrial matrix for translocation and folding of precursor proteins.
    Nature. 1990 Nov 8;348(6297):137-43 PMID: 2234077
  16. Internal ATP is the only energy requirement for the translocation of precursor proteins across chloroplastic membranes.
    J Biol Chem. 1989 Apr 25;264(12):6730-6 PMID: 2708340
  17. S. cerevisiae encodes an essential protein homologous in sequence and function to mammalian BiP.
    Cell. 1989 Jun 30;57(7):1223-36 PMID: 2661019
  18. Translocation of ATP into the lumen of rough endoplasmic reticulum-derived vesicles and its binding to luminal proteins including BiP (GRP 78) and GRP 94.
    J Biol Chem. 1992 Feb 25;267(6):3983-90 PMID: 1740446
  19. Binding protein BiP is required for translocation of secretory proteins into the endoplasmic reticulum in Saccharomyces cerevisiae.
    Proc Natl Acad Sci U S A. 1991 Feb 15;88(4):1565-9 PMID: 1996357
  20. Functional messenger RNAs are produced by SP6 in vitro transcription of cloned cDNAs.
    Nucleic Acids Res. 1984 Sep 25;12(18):7057-70 PMID: 6207484
  21. Secretion in yeast: reconstitution of the translocation and glycosylation of alpha-factor and invertase in a homologous cell-free system.
    Cell. 1986 Feb 28;44(4):619-28 PMID: 3512097
  22. Secretory protein translocation in a yeast cell-free system can occur posttranslationally and requires ATP hydrolysis.
    J Cell Biol. 1986 May;102(5):1543-50 PMID: 3517001
  23. 8-azido-ADP, a covalent-binding inhibitor of mitochondrial adenine nucleotide translocation.
    FEBS Lett. 1976 Apr 15;64(1):185-9 PMID: 131709
  24. Defective co-translational formation of disulphide bonds in protein disulphide-isomerase-deficient microsomes.
    Nature. 1988 Oct 13;335(6191):649-51 PMID: 3173483
  25. Transport and assembly processes in the endoplasmic reticulum.
    Semin Cell Biol. 1990 Feb;1(1):65-72 PMID: 1983272
  26. A subfamily of stress proteins facilitates translocation of secretory and mitochondrial precursor polypeptides.
    Nature. 1988 Apr 28;332(6167):800-5 PMID: 3282178
  27. Translocation of proteins across the mitochondrial inner membrane, but not into the outer membrane, requires nucleoside triphosphates in the matrix.
    Proc Natl Acad Sci U S A. 1989 Nov;86(21):8432-6 PMID: 2554316
  28. Protein transport across the ER membrane.
    Trends Biochem Sci. 1990 Sep;15(9):355-8 PMID: 2238046
  29. An ATP-binding membrane protein is required for protein translocation across the endoplasmic reticulum membrane.
    Cell Regul. 1991 Oct;2(10):851-9 PMID: 1801920
  30. 70K heat shock related proteins stimulate protein translocation into microsomes.
    Nature. 1988 Apr 28;332(6167):805-10 PMID: 3282179
  31. Secretion in yeast. Purification and in vitro translocation of chemical amounts of prepro-alpha-factor.
    J Biol Chem. 1991 Jul 25;266(21):13811-4 PMID: 1856213
  32. Loss of BiP/GRP78 function blocks translocation of secretory proteins in yeast.
    J Cell Biol. 1990 Jun;110(6):1885-95 PMID: 2190988
  33. Exploring the adenine nucleotide binding sites on mitochondrial F1-ATPase with a new photoaffinity probe, 3'-O-(4-benzoyl)benzoyl adenosine 5'-triphosphate.
    J Biol Chem. 1982 Mar 25;257(6):2834-41 PMID: 6460764
  34. Molecular aspects of band 3 protein-mediated anion transport across the red blood cell membrane.
    Rev Physiol Biochem Pharmacol. 1986;103:61-203 PMID: 2421388
  35. Characterization of the solubilization of lipid bilayers by surfactants.
    Biochim Biophys Acta. 1985 Dec 19;821(3):470-8 PMID: 4074739
  36. The ADP/ATP carrier from yeast (AAC-2) is uniquely suited for the assignment of the binding center by photoaffinity labeling.
    FEBS Lett. 1989 Feb 27;244(2):421-6 PMID: 2537764
  37. A precursor protein partly translocated into yeast mitochondria is bound to a 70 kd mitochondrial stress protein.
    EMBO J. 1990 Dec;9(13):4315-22 PMID: 2265609
  38. Mutants in three novel complementation groups inhibit membrane protein insertion into and soluble protein translocation across the endoplasmic reticulum membrane of Saccharomyces cerevisiae.
    J Cell Biol. 1992 Feb;116(3):597-604 PMID: 1730771
  39. In vitro protein translocation across the yeast endoplasmic reticulum: ATP-dependent posttranslational translocation of the prepro-alpha-factor.
    Cell. 1986 May 9;45(3):397-406 PMID: 3009026
  40. KAR2, a karyogamy gene, is the yeast homolog of the mammalian BiP/GRP78 gene.
    Cell. 1989 Jun 30;57(7):1211-21 PMID: 2661018
  41. Membrane proteins related to anion permeability of human red blood cells. I. Localization of disulfonic stilbene binding sites in proteins involved in permeation.
    J Membr Biol. 1974;15(3):207-26 PMID: 4838037
  42. Probing the active site of the reconstituted aspartate/glutamate carrier from bovine heart mitochondria: carbodiimide-catalyzed acylation of a functional lysine residue.
    Biochim Biophys Acta. 1992 Jan 10;1103(1):13-24 PMID: 1346091
Article Info
Journal
The EMBO journal
Abbr.
EMBO J
ISSN
0261-4189
Published
1993-02-00
Pages
659-66
Language
English
Region
England
NLM ID
8208664
PMCID
PMC413250
Subset
IM
External Links
PubMed source PMC Full Text View journal details
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