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Volumn 1480, Issue 1-2, 2000, Pages 267-277

Domain-specific spectroscopy of 5-hydroxytryptophan-containing variants of Escherichia coli DnaJ

Author keywords

Fluorescence; Heat shock protein; Protein folding; Tryptophan analog

Indexed keywords

5 HYDROXYTRYPTOPHAN; HEAT SHOCK PROTEIN; TRYPTOPHAN;

EID: 0034647654     PISSN: 01674838     EISSN: None     Source Type: Journal    
DOI: 10.1016/S0167-4838(00)00078-9     Document Type: Article
Times cited : (3)

References (49)
  • 2
    • 0028281387 scopus 로고
    • Paralysis and early death in cysteine string protein mutants of Drosophila
    • Zinsmaier K.E., Eberle K.K., Buchner E., Walter N., Benzer S. Paralysis and early death in cysteine string protein mutants of Drosophila. Science. 263:1994;977-980.
    • (1994) Science , vol.263 , pp. 977-980
    • Zinsmaier, K.E.1    Eberle, K.K.2    Buchner, E.3    Walter, N.4    Benzer, S.5
  • 3
    • 0027102871 scopus 로고
    • YDJ1p facilitates polypeptide translocation across different intracellular membranes by a conserved mechanism
    • Caplan A.J., Cyr D.M., Douglas M.G. YDJ1p facilitates polypeptide translocation across different intracellular membranes by a conserved mechanism. Cell. 71:1992;1143-1155.
    • (1992) Cell , vol.71 , pp. 1143-1155
    • Caplan, A.J.1    Cyr, D.M.2    Douglas, M.G.3
  • 4
    • 0025976271 scopus 로고
    • Heat-shock proteins can substitute for SecB function during protein export in Escherichia coli
    • Altman E., Kumamoto C.A., Emr S.D. Heat-shock proteins can substitute for SecB function during protein export in Escherichia coli. EMBO J. 10:1991;239-245.
    • (1991) EMBO J. , vol.10 , pp. 239-245
    • Altman, E.1    Kumamoto, C.A.2    Emr, S.D.3
  • 5
    • 0026644011 scopus 로고
    • DnaK and DnaJ heat shock proteins participate in protein export in Escherichia coli
    • Wild J., Altman E., Yura T., Gross C.A. DnaK and DnaJ heat shock proteins participate in protein export in Escherichia coli. Genes Dev. 6:1992;1165-1172.
    • (1992) Genes Dev. , vol.6 , pp. 1165-1172
    • Wild, J.1    Altman, E.2    Yura, T.3    Gross, C.A.4
  • 6
    • 0026569763 scopus 로고
    • MAS5, a yeast homolog of DnaJ involved in mitochondrial protein import
    • Atencio D.P., Yaffe M.P. MAS5, a yeast homolog of DnaJ involved in mitochondrial protein import. Mol. Cell. Biol. 12:1992;283-291.
    • (1992) Mol. Cell. Biol. , vol.12 , pp. 283-291
    • Atencio, D.P.1    Yaffe, M.P.2
  • 7
    • 0026596223 scopus 로고
    • Successive action of DnaK, DnaJ and GroEL along the pathway of chaperone-mediated protein folding
    • Langer T., Lu C., Echols H., Flanagan J., Hayer M.K., Hartl F.U. Successive action of DnaK, DnaJ and GroEL along the pathway of chaperone-mediated protein folding. Nature. 356:1992;683-689.
    • (1992) Nature , vol.356 , pp. 683-689
    • Langer, T.1    Lu, C.2    Echols, H.3    Flanagan, J.4    Hayer, M.K.5    Hartl, F.U.6
  • 8
    • 0030030946 scopus 로고    scopus 로고
    • A zinc finger-like domain of the molecular chaperone DnaJ is involved in binding to denatured protein substrates
    • Szabo A., Korszun R., Hartl F.-U., Flanagan J. A zinc finger-like domain of the molecular chaperone DnaJ is involved in binding to denatured protein substrates. EMBO J. 15:1996;408-417.
    • (1996) EMBO J. , vol.15 , pp. 408-417
    • Szabo, A.1    Korszun, R.2    Hartl, F.-U.3    Flanagan, J.4
  • 9
    • 0027425585 scopus 로고
    • Control of folding and membrane translocation by binding of the chaperone DnaJ to nascent polypeptides
    • Hendrick J.P., Langer T., Davis T.A., Hartl F.U., Wiedmann M. Control of folding and membrane translocation by binding of the chaperone DnaJ to nascent polypeptides. Proc. Natl. Acad. Sci. USA. 90:1993;10216-10220.
    • (1993) Proc. Natl. Acad. Sci. USA , vol.90 , pp. 10216-10220
    • Hendrick, J.P.1    Langer, T.2    Davis, T.A.3    Hartl, F.U.4    Wiedmann, M.5
  • 10
    • 0025266168 scopus 로고
    • Three Escherichia coli heat shock proteins are required for P1 plasmid DNA replication: Formation of an active complex between E. coli DnaJ protein and the P1 initiator protein
    • Wickner S.H. Three Escherichia coli heat shock proteins are required for P1 plasmid DNA replication: formation of an active complex between E. coli DnaJ protein and the P1 initiator protein. Proc. Natl. Acad. Sci. USA. 87:1990;2690-2694.
    • (1990) Proc. Natl. Acad. Sci. USA , vol.87 , pp. 2690-2694
    • Wickner, S.H.1
  • 11
    • 0025248655 scopus 로고
    • Physical interactions between bacteriophage and Escherichia coli proteins required for initiation of lambda DNA replication
    • Liberek K., Osipiuk J., Zylicz M., Ang D., Skorko J., Georgopoulos C. Physical interactions between bacteriophage and Escherichia coli proteins required for initiation of lambda DNA replication. J. Biol. Chem. 265:1990;3022-3029.
    • (1990) J. Biol. Chem. , vol.265 , pp. 3022-3029
    • Liberek, K.1    Osipiuk, J.2    Zylicz, M.3    Ang, D.4    Skorko, J.5    Georgopoulos, C.6
  • 12
    • 0028353336 scopus 로고
    • DnaJ-like proteins: Molecular chaperones and specific regulators of hsp70
    • Cyr D.M., Langer T., Douglas M.G. DnaJ-like proteins: molecular chaperones and specific regulators of hsp70. Trends Biochem. Sci. 19:1994;176-181.
    • (1994) Trends Biochem. Sci. , vol.19 , pp. 176-181
    • Cyr, D.M.1    Langer, T.2    Douglas, M.G.3
  • 13
    • 0025875276 scopus 로고
    • Function of DnaJ and DnaK as chaperones in origin-specific DNA binding by RepA
    • Wickner S., Hoskins J., McKenney K. Function of DnaJ and DnaK as chaperones in origin-specific DNA binding by RepA. Nature. 350:1991;165-167.
    • (1991) Nature , vol.350 , pp. 165-167
    • Wickner, S.1    Hoskins, J.2    McKenney, K.3
  • 14
    • 0029094250 scopus 로고
    • Divergent effects of ATP on the binding of the DnaK and DnaJ chaperones to each other, or to their various native and denatured protein substrates
    • Wawrzynow A., Zylicz M. Divergent effects of ATP on the binding of the DnaK and DnaJ chaperones to each other, or to their various native and denatured protein substrates. J. Biol. Chem. 270:1995;19300-19306.
    • (1995) J. Biol. Chem. , vol.270 , pp. 19300-19306
    • Wawrzynow, A.1    Zylicz, M.2
  • 15
    • 0026059137 scopus 로고
    • Peptide-binding specificity of the molecular chaperone Bip
    • Flynn G.C., Pohl J., Flocco M.T., Rothman J.E. Peptide-binding specificity of the molecular chaperone Bip. Nature. 353:1991;726-730.
    • (1991) Nature , vol.353 , pp. 726-730
    • Flynn, G.C.1    Pohl, J.2    Flocco, M.T.3    Rothman, J.E.4
  • 16
    • 0026595140 scopus 로고
    • Different conformations for the same polypeptide bound to chaperones DnaK and GroEL
    • Landry S.J., Jordan R., McMacken R., Gierasch L.M. Different conformations for the same polypeptide bound to chaperones DnaK and GroEL. Nature. 355:1992;455-457.
    • (1992) Nature , vol.355 , pp. 455-457
    • Landry, S.J.1    Jordan, R.2    McMacken, R.3    Gierasch, L.M.4
  • 18
    • 0026696625 scopus 로고
    • Physical interaction between heat shock proteins DnaK, DnaJ, and GrpE and the bacterial heat shock transcription factor-sigma(32)
    • Gamer J., Bujard H., Bukau B. Physical interaction between heat shock proteins DnaK, DnaJ, and GrpE and the bacterial heat shock transcription factor-sigma(32). Cell. 69:1992;833-842.
    • (1992) Cell , vol.69 , pp. 833-842
    • Gamer, J.1    Bujard, H.2    Bukau, B.3
  • 19
    • 0024978379 scopus 로고
    • Ordered assembly of nucleoprotein structures at the bacteriophage lambda replication origin during the initiation of DNA replication
    • Alfano C., McMacken R. Ordered assembly of nucleoprotein structures at the bacteriophage lambda replication origin during the initiation of DNA replication. J. Biol. Chem. 264:1989;10699-10708.
    • (1989) J. Biol. Chem. , vol.264 , pp. 10699-10708
    • Alfano, C.1    McMacken, R.2
  • 20
    • 0027427986 scopus 로고
    • DnaK, DnaJ and GrpE form a cellular chaperone machinery capable of repairing heat-induced protein damage
    • Schroder H., Langer T., Hartl F.U., Bukau B. DnaK, DnaJ and GrpE form a cellular chaperone machinery capable of repairing heat-induced protein damage. EMBO J. 12:1993;4137-4144.
    • (1993) EMBO J. , vol.12 , pp. 4137-4144
    • Schroder, H.1    Langer, T.2    Hartl, F.U.3    Bukau, B.4
  • 21
    • 0026690820 scopus 로고
    • Topology and functional domains of Sec63p, an endoplasmic reticulum membrane protein required for secretory protein translocation
    • Feldheim D., Rothblatt J., Schekman R. Topology and functional domains of Sec63p, an endoplasmic reticulum membrane protein required for secretory protein translocation. Mol. Cell. Biol. 12:1992;3288-3296.
    • (1992) Mol. Cell. Biol. , vol.12 , pp. 3288-3296
    • Feldheim, D.1    Rothblatt, J.2    Schekman, R.3
  • 22
    • 0028170215 scopus 로고
    • 2-terminal 108 amino acids of the Escherichia coli DnaJ protein stimulate the ATPase activity of DnaK and are sufficient for lambda replication
    • 2-terminal 108 amino acids of the Escherichia coli DnaJ protein stimulate the ATPase activity of DnaK and are sufficient for lambda replication. J. Biol. Chem. 269:1994;5446-5451.
    • (1994) J. Biol. Chem. , vol.269 , pp. 5446-5451
    • Wall, D.1    Zylicz, M.2    Georgopoulos, C.3
  • 23
    • 0032568541 scopus 로고    scopus 로고
    • Role of the J-domain in the cooperation of Hsp40 with Hsp70
    • Greene M.K., Maskos K., Landry S.J. Role of the J-domain in the cooperation of Hsp40 with Hsp70. Proc. Natl. Acad. Sci. USA. 95:1998;6108-6113.
    • (1998) Proc. Natl. Acad. Sci. USA , vol.95 , pp. 6108-6113
    • Greene, M.K.1    Maskos, K.2    Landry, S.J.3
  • 24
    • 0027987996 scopus 로고
    • NMR structure determination of the Escherichia coli DnaJ molecular chaperone: Secondary structure and backbone fold of the N-terminal region (residues 2-108) containing the highly conserved J domain
    • Szyperski T., Pellecchia M., Wall D., Georgopoulos C., Wuthrich K. NMR structure determination of the Escherichia coli DnaJ molecular chaperone: secondary structure and backbone fold of the N-terminal region (residues 2-108) containing the highly conserved J domain. Proc. Natl. Acad. Sci. USA. 91:1994;11343-11347.
    • (1994) Proc. Natl. Acad. Sci. USA , vol.91 , pp. 11343-11347
    • Szyperski, T.1    Pellecchia, M.2    Wall, D.3    Georgopoulos, C.4    Wuthrich, K.5
  • 25
    • 15844372190 scopus 로고    scopus 로고
    • A bipartite signaling mechanism involved in DnaJ-mediated activation of the Escherichia coli DnaK protein
    • Karzai A.W., McMacken R. A bipartite signaling mechanism involved in DnaJ-mediated activation of the Escherichia coli DnaK protein. J. Biol. Chem. 271:1996;11236-11246.
    • (1996) J. Biol. Chem. , vol.271 , pp. 11236-11246
    • Karzai, A.W.1    McMacken, R.2
  • 26
    • 0027169533 scopus 로고
    • Eukaryotic DnaJ homologs and the specificity of hsp70 activity
    • Silver P.A., Way J.C. Eukaryotic DnaJ homologs and the specificity of hsp70 activity. Cell. 74:1993;5-6.
    • (1993) Cell , vol.74 , pp. 5-6
    • Silver, P.A.1    Way, J.C.2
  • 29
    • 0026586906 scopus 로고
    • A module of the DnaJ heat shock proteins found in malaria parasites
    • Bork P., Sander C., Valencia A., Bukau B. A module of the DnaJ heat shock proteins found in malaria parasites. Trends Biochem. Sci. 17:1992;129.
    • (1992) Trends Biochem. Sci. , vol.17 , pp. 129
    • Bork, P.1    Sander, C.2    Valencia, A.3    Bukau, B.4
  • 30
    • 0027378450 scopus 로고
    • Resolution of the fluorescence equilibrium unfolding profile of trp aporepressor using single tryptophan mutants
    • Royer C.A., Mann C.J., Matthews C.R. Resolution of the fluorescence equilibrium unfolding profile of trp aporepressor using single tryptophan mutants. Protein Sci. 2:1993;1844-1852.
    • (1993) Protein Sci. , vol.2 , pp. 1844-1852
    • Royer, C.A.1    Mann, C.J.2    Matthews, C.R.3
  • 31
    • 0027438393 scopus 로고
    • Tryptophan replacements in the trp aporepressor from Escherichia coli: Probing the equilibrium and kinetic folding models
    • Mann C.J., Royer C.A., Matthews C.R. Tryptophan replacements in the trp aporepressor from Escherichia coli: probing the equilibrium and kinetic folding models. Protein Sci. 2:1993;1853-1861.
    • (1993) Protein Sci. , vol.2 , pp. 1853-1861
    • Mann, C.J.1    Royer, C.A.2    Matthews, C.R.3
  • 32
    • 0028966987 scopus 로고
    • Contribution of individual tryptophan residues to the fluorescence spectrum of native and denatured forms of human carbonic anhydrase II
    • Martensson L.-G., Jonasson P., Freskgard P.-O., Svensson M., Carlsson U., Jonsson B.-H. Contribution of individual tryptophan residues to the fluorescence spectrum of native and denatured forms of human carbonic anhydrase II. Biochemistry. 34:1995;1011-1021.
    • (1995) Biochemistry , vol.34 , pp. 1011-1021
    • Martensson, L.-G.1    Jonasson, P.2    Freskgard, P.-O.3    Svensson, M.4    Carlsson, U.5    Jonsson, B.-H.6
  • 33
    • 0019883167 scopus 로고
    • Tryptophanyl fluorescence heterogeneity of apomyoglobins: Correlation with the presence of two distinct structural domains
    • Irace G., Balestrieri C., Parlato G., Servillo L., Colonna G. Tryptophanyl fluorescence heterogeneity of apomyoglobins: correlation with the presence of two distinct structural domains. Biochemistry. 20:1981;792-799.
    • (1981) Biochemistry , vol.20 , pp. 792-799
    • Irace, G.1    Balestrieri, C.2    Parlato, G.3    Servillo, L.4    Colonna, G.5
  • 34
    • 0029865110 scopus 로고    scopus 로고
    • Transcriptional activation domain of the Herpesvirus protein VP16 becomes conformationally constrained upon interaction with basal transcription factors
    • Shen F., Triezenberg S.J., Hensley P., Porter D., Knutson J.R. Transcriptional activation domain of the Herpesvirus protein VP16 becomes conformationally constrained upon interaction with basal transcription factors. J. Biol. Chem. 271:1996;4827-4837.
    • (1996) J. Biol. Chem. , vol.271 , pp. 4827-4837
    • Shen, F.1    Triezenberg, S.J.2    Hensley, P.3    Porter, D.4    Knutson, J.R.5
  • 35
    • 0027537906 scopus 로고
    • 5-Hydroxytryptophan as a new intrinsic probe for investigating protein DNA interactions by analytical ultracentrifugation: Study of the effect of DNA on self-assembly of the bacteriophage lambda cI repressor
    • Laue T.M., Senear D.F., Eaton S., Ross J.B.A. 5-Hydroxytryptophan as a new intrinsic probe for investigating protein DNA interactions by analytical ultracentrifugation: study of the effect of DNA on self-assembly of the bacteriophage lambda cI repressor. Biochemistry. 32:1993;2469-2472.
    • (1993) Biochemistry , vol.32 , pp. 2469-2472
    • Laue, T.M.1    Senear, D.F.2    Eaton, S.3    Ross, J.B.A.4
  • 36
    • 0032560613 scopus 로고    scopus 로고
    • Incorporation of tryptophan analogues into staphylococcal nuclease: Stability toward thermal and guanidine-HCl induced unfolding
    • Wong C.-Y., Eftink M.R. Incorporation of tryptophan analogues into staphylococcal nuclease: stability toward thermal and guanidine-HCl induced unfolding. Biochemistry. 37:1998;8947-8953.
    • (1998) Biochemistry , vol.37 , pp. 8947-8953
    • Wong, C.-Y.1    Eftink, M.R.2
  • 37
    • 0030935256 scopus 로고    scopus 로고
    • The T/t common exon of simian virus 40, JC, and BK polyomavirus T antigens can functionally replace the J-domain of the Escherichia coli DnaJ molecular chaperone
    • Kelley W.L., Georgopoulos C. The T/t common exon of simian virus 40, JC, and BK polyomavirus T antigens can functionally replace the J-domain of the Escherichia coli DnaJ molecular chaperone. Proc. Natl. Acad. Sci. USA. 94:1997;3679-3684.
    • (1997) Proc. Natl. Acad. Sci. USA , vol.94 , pp. 3679-3684
    • Kelley, W.L.1    Georgopoulos, C.2
  • 38
    • 0024412948 scopus 로고
    • Nuclear magnetic resonance studies of 6-fluorotryptophan-substituted rat cellular retinol-binding protein II produced in Escherichia coli
    • Li E., Qian S.-j., Nader L., Yang N.-c.C., d'Avignon A., Sacchettini J.C., Gordon J.I. Nuclear magnetic resonance studies of 6-fluorotryptophan-substituted rat cellular retinol-binding protein II produced in Escherichia coli. J. Biol. Chem. 264:1989;17041-17048.
    • (1989) J. Biol. Chem. , vol.264 , pp. 17041-17048
    • Li, E.1    Qian, S.-J.2    Nader, L.3    Yang, N.-C.C.4    D'Avignon, A.5    Sacchettini, J.C.6    Gordon, J.I.7
  • 39
    • 0025420076 scopus 로고
    • Measuring and increasing protein stability
    • Pace C.N. Measuring and increasing protein stability. Trends Biotechnol. 8:1990;93-98.
    • (1990) Trends Biotechnol. , vol.8 , pp. 93-98
    • Pace, C.N.1
  • 40
    • 84908824031 scopus 로고
    • Methods of polyacrylamide gel electrophoresis in the analysis and preparation of plant polypeptides
    • in: M. Edelman, R.B. Hallick, N.-H. Chua (Eds.), Elsevier Biomedical Press, New York
    • R. Piccioni, G. Bellemare, N.-H. Chua, Methods of polyacrylamide gel electrophoresis in the analysis and preparation of plant polypeptides in: M. Edelman, R.B. Hallick, N.-H. Chua (Eds.), Methods in Chloroplast Molecular Biology, Elsevier Biomedical Press, New York, 1982, pp. 985-1014.
    • (1982) Methods in Chloroplast Molecular Biology , pp. 985-1014
    • Piccioni, R.1    Bellemare, G.2    Chua, N.-H.3
  • 41
    • 0026244229 scopus 로고
    • MOLSCRIPT: A Program to produce both detailed and schematic plots of protein structures
    • Kraulis P.J. MOLSCRIPT: A Program to produce both detailed and schematic plots of protein structures. J. Appl. Crystallogr. 24:1991;946-950.
    • (1991) J. Appl. Crystallogr. , vol.24 , pp. 946-950
    • Kraulis, P.J.1
  • 42
    • 0015230409 scopus 로고
    • Solute perturbation of protein fluorescence: The quenching of the tryptophanyl fluorescence of model compounds and of lysozyme by iodide ion
    • Lehrer S.S. Solute perturbation of protein fluorescence: the quenching of the tryptophanyl fluorescence of model compounds and of lysozyme by iodide ion. Biochemistry. 10:1971;3254-3263.
    • (1971) Biochemistry , vol.10 , pp. 3254-3263
    • Lehrer, S.S.1
  • 43
    • 0030581175 scopus 로고    scopus 로고
    • NMR structure of the J-domain and the gly/phe-rich region of the Escherichia coli DnaJ chaperone
    • Pellecchia M., Szyperski T., Wall D., Georgopoulos C., Wuthrich K. NMR structure of the J-domain and the gly/phe-rich region of the Escherichia coli DnaJ chaperone. J. Mol. Biol. 260:1996;236-250.
    • (1996) J. Mol. Biol. , vol.260 , pp. 236-250
    • Pellecchia, M.1    Szyperski, T.2    Wall, D.3    Georgopoulos, C.4    Wuthrich, K.5
  • 45
    • 0026787837 scopus 로고
    • Use of protein unfolding studies to determine the conformational and dimeric stabilities of HIV-1 and SIV proteases
    • Grant S.K., Deckman I.C., Culp J.S., Minnich M.D., Brooks I.S., Hensley P., Debouck C., Meek T.D. Use of protein unfolding studies to determine the conformational and dimeric stabilities of HIV-1 and SIV proteases. Biochemistry. 31:1992;9491-9501.
    • (1992) Biochemistry , vol.31 , pp. 9491-9501
    • Grant, S.K.1    Deckman, I.C.2    Culp, J.S.3    Minnich, M.D.4    Brooks, I.S.5    Hensley, P.6    Debouck, C.7    Meek, T.D.8
  • 46
    • 0026086650 scopus 로고
    • Equilibrium unfolding of class pi glutathione s-transferase
    • Dirr H.W., Reinemer P. Equilibrium unfolding of class pi glutathione s-transferase. Biochem. Biophys. Res. Commun. 180:1991;294-300.
    • (1991) Biochem. Biophys. Res. Commun. , vol.180 , pp. 294-300
    • Dirr, H.W.1    Reinemer, P.2
  • 47
    • 0030980794 scopus 로고    scopus 로고
    • Enhancement of protein spectra with tryptophan analogs: Fluorescence spectroscopy of protein-protein and protein-nucleic interactions
    • Ross J.B., Szabo A.G., Hogue C.W. Enhancement of protein spectra with tryptophan analogs: fluorescence spectroscopy of protein-protein and protein-nucleic interactions. Methods Enzymol. 278:1997;151-190.
    • (1997) Methods Enzymol. , vol.278 , pp. 151-190
    • Ross, J.B.1    Szabo, A.G.2    Hogue, C.W.3
  • 48
    • 0030446234 scopus 로고    scopus 로고
    • Oligomers of the cytoplasmic fragment from the Escherichia coli aspartate receptor dissociate through an unfolded transition state
    • Seely S.K., Wittrock G.K., Thompson L.K., Weis R.M. Oligomers of the cytoplasmic fragment from the Escherichia coli aspartate receptor dissociate through an unfolded transition state. Biochemistry. 35:1996;16336-16345.
    • (1996) Biochemistry , vol.35 , pp. 16336-16345
    • Seely, S.K.1    Wittrock, G.K.2    Thompson, L.K.3    Weis, R.M.4
  • 49
    • 0029081273 scopus 로고
    • Urea-induced dissociation and unfolding of dodecameric glutamine synthetase from Escherichia coli: Calorimetric and spectral studies
    • Zolkiewski M., Nosworthy N.J., Ginsburg A. Urea-induced dissociation and unfolding of dodecameric glutamine synthetase from Escherichia coli: calorimetric and spectral studies. Protein Sci. 4:1995;1544-1552.
    • (1995) Protein Sci. , vol.4 , pp. 1544-1552
    • Zolkiewski, M.1    Nosworthy, N.J.2    Ginsburg, A.3


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