Role of the ATP synthase α-subunit in conferring sensitivity to tentoxin

W. C. Tucker, Ziyun Du, R. Hein, Z. Gromet-Elhanan, M. L. Richter

Research output: Contribution to journalArticle

16 Citations (Scopus)

Abstract

Tentoxin, produced by phytopathogenic fungi, selectively affects the function of the ATP synthase enzymes of certain sensitive plant species. Binding of tentoxin to a high affinity (Ki ≈ 10 nM) site on the chloroplast F1 (CF1) strongly inhibits catalytic function, whereas binding to a second, lower affinity site (Kd > 10 μM) leads to restoration and even stimulation of catalytic activity. Sensitivity to tentoxin has been shown to be due, in part, to the nature of the amino acid residue at position 83 on the catalytic β subunit of CF1. An aspartate in this position is required, but is not sufficient, for tentoxin inhibition. By comparison with the solved structure of mitochondrial F1 [Abrahams, J. P., Leslie, A. G. W., Lutter, R., and Walker, J. E. (1994) Nature 370, 621-628], Asp83 is probably located at an interface between α and β subunits on CF1 where residues on the α subunit could also participate in tentoxin binding. A hybrid core F1 enzyme assembled with β and γ subunits of the tentoxin-sensitive spinach CF1, and an α subunit of the tentoxin-insensitive photosynthetic bacterium Rhodospirillum rubrum F1 (RrF1), was stimulated but not inhibited by tentoxin [Tucker, W. C., Du, Z., Gromet-Elhanan, Z. and Richter, M. L. (2001) Eur. J. Biochem. 268, 2179-2186]. In this study, chimeric α subunits were prepared by introducing short segments of the spinach CF1 α subunit from a poorly conserved region which is immediately adjacent to β-Asp83 in the crystal structure, into equivalent positions in the RrF1 α subunit using oligonucleotide-directed mutagenesis. Hybrid enzymes containing these chimeric α subunits had both the high affinity inhibitory tentoxin binding site and the lower affinity stimulatory site. Changing β-Asp83 to leucine resulted in loss of both inhibition and stimulation by tentoxin in the chimeras. The results indicate that tentoxin inhibition requires additional α residues that are not present on the RrF1 α subunit. A structural model of a putative inhibitory tentoxin binding pocket is presented.

Original languageEnglish (US)
Pages (from-to)7542-7548
Number of pages7
JournalBiochemistry
Volume40
Issue number25
DOIs
StatePublished - Jun 26 2001

Fingerprint

Adenosine Triphosphate
Chloroplasts
Rhodospirillum rubrum
Spinacia oleracea
tentoxin
Enzymes
Mutagenesis
Structural Models
Site-Directed Mutagenesis
Fungi
Aspartic Acid
Leucine
Oligonucleotides
Restoration
Catalyst activity
Catalytic Domain
Bacteria
Crystal structure
Binding Sites
Amino Acids

All Science Journal Classification (ASJC) codes

  • Biochemistry

Cite this

Tucker, W. C., Du, Z., Hein, R., Gromet-Elhanan, Z., & Richter, M. L. (2001). Role of the ATP synthase α-subunit in conferring sensitivity to tentoxin. Biochemistry, 40(25), 7542-7548. https://doi.org/10.1021/bi0105227

Role of the ATP synthase α-subunit in conferring sensitivity to tentoxin. / Tucker, W. C.; Du, Ziyun; Hein, R.; Gromet-Elhanan, Z.; Richter, M. L.

In: Biochemistry, Vol. 40, No. 25, 26.06.2001, p. 7542-7548.

Research output: Contribution to journalArticle

Tucker, WC, Du, Z, Hein, R, Gromet-Elhanan, Z & Richter, ML 2001, 'Role of the ATP synthase α-subunit in conferring sensitivity to tentoxin', Biochemistry, vol. 40, no. 25, pp. 7542-7548. https://doi.org/10.1021/bi0105227
Tucker WC, Du Z, Hein R, Gromet-Elhanan Z, Richter ML. Role of the ATP synthase α-subunit in conferring sensitivity to tentoxin. Biochemistry. 2001 Jun 26;40(25):7542-7548. https://doi.org/10.1021/bi0105227
Tucker, W. C. ; Du, Ziyun ; Hein, R. ; Gromet-Elhanan, Z. ; Richter, M. L. / Role of the ATP synthase α-subunit in conferring sensitivity to tentoxin. In: Biochemistry. 2001 ; Vol. 40, No. 25. pp. 7542-7548.
@article{99c69b1ed4444e8f950b9799a2e6cb71,
title = "Role of the ATP synthase α-subunit in conferring sensitivity to tentoxin",
abstract = "Tentoxin, produced by phytopathogenic fungi, selectively affects the function of the ATP synthase enzymes of certain sensitive plant species. Binding of tentoxin to a high affinity (Ki ≈ 10 nM) site on the chloroplast F1 (CF1) strongly inhibits catalytic function, whereas binding to a second, lower affinity site (Kd > 10 μM) leads to restoration and even stimulation of catalytic activity. Sensitivity to tentoxin has been shown to be due, in part, to the nature of the amino acid residue at position 83 on the catalytic β subunit of CF1. An aspartate in this position is required, but is not sufficient, for tentoxin inhibition. By comparison with the solved structure of mitochondrial F1 [Abrahams, J. P., Leslie, A. G. W., Lutter, R., and Walker, J. E. (1994) Nature 370, 621-628], Asp83 is probably located at an interface between α and β subunits on CF1 where residues on the α subunit could also participate in tentoxin binding. A hybrid core F1 enzyme assembled with β and γ subunits of the tentoxin-sensitive spinach CF1, and an α subunit of the tentoxin-insensitive photosynthetic bacterium Rhodospirillum rubrum F1 (RrF1), was stimulated but not inhibited by tentoxin [Tucker, W. C., Du, Z., Gromet-Elhanan, Z. and Richter, M. L. (2001) Eur. J. Biochem. 268, 2179-2186]. In this study, chimeric α subunits were prepared by introducing short segments of the spinach CF1 α subunit from a poorly conserved region which is immediately adjacent to β-Asp83 in the crystal structure, into equivalent positions in the RrF1 α subunit using oligonucleotide-directed mutagenesis. Hybrid enzymes containing these chimeric α subunits had both the high affinity inhibitory tentoxin binding site and the lower affinity stimulatory site. Changing β-Asp83 to leucine resulted in loss of both inhibition and stimulation by tentoxin in the chimeras. The results indicate that tentoxin inhibition requires additional α residues that are not present on the RrF1 α subunit. A structural model of a putative inhibitory tentoxin binding pocket is presented.",
author = "Tucker, {W. C.} and Ziyun Du and R. Hein and Z. Gromet-Elhanan and Richter, {M. L.}",
year = "2001",
month = "6",
day = "26",
doi = "10.1021/bi0105227",
language = "English (US)",
volume = "40",
pages = "7542--7548",
journal = "Biochemistry",
issn = "0006-2960",
publisher = "American Chemical Society",
number = "25",

}

TY - JOUR

T1 - Role of the ATP synthase α-subunit in conferring sensitivity to tentoxin

AU - Tucker, W. C.

AU - Du, Ziyun

AU - Hein, R.

AU - Gromet-Elhanan, Z.

AU - Richter, M. L.

PY - 2001/6/26

Y1 - 2001/6/26

N2 - Tentoxin, produced by phytopathogenic fungi, selectively affects the function of the ATP synthase enzymes of certain sensitive plant species. Binding of tentoxin to a high affinity (Ki ≈ 10 nM) site on the chloroplast F1 (CF1) strongly inhibits catalytic function, whereas binding to a second, lower affinity site (Kd > 10 μM) leads to restoration and even stimulation of catalytic activity. Sensitivity to tentoxin has been shown to be due, in part, to the nature of the amino acid residue at position 83 on the catalytic β subunit of CF1. An aspartate in this position is required, but is not sufficient, for tentoxin inhibition. By comparison with the solved structure of mitochondrial F1 [Abrahams, J. P., Leslie, A. G. W., Lutter, R., and Walker, J. E. (1994) Nature 370, 621-628], Asp83 is probably located at an interface between α and β subunits on CF1 where residues on the α subunit could also participate in tentoxin binding. A hybrid core F1 enzyme assembled with β and γ subunits of the tentoxin-sensitive spinach CF1, and an α subunit of the tentoxin-insensitive photosynthetic bacterium Rhodospirillum rubrum F1 (RrF1), was stimulated but not inhibited by tentoxin [Tucker, W. C., Du, Z., Gromet-Elhanan, Z. and Richter, M. L. (2001) Eur. J. Biochem. 268, 2179-2186]. In this study, chimeric α subunits were prepared by introducing short segments of the spinach CF1 α subunit from a poorly conserved region which is immediately adjacent to β-Asp83 in the crystal structure, into equivalent positions in the RrF1 α subunit using oligonucleotide-directed mutagenesis. Hybrid enzymes containing these chimeric α subunits had both the high affinity inhibitory tentoxin binding site and the lower affinity stimulatory site. Changing β-Asp83 to leucine resulted in loss of both inhibition and stimulation by tentoxin in the chimeras. The results indicate that tentoxin inhibition requires additional α residues that are not present on the RrF1 α subunit. A structural model of a putative inhibitory tentoxin binding pocket is presented.

AB - Tentoxin, produced by phytopathogenic fungi, selectively affects the function of the ATP synthase enzymes of certain sensitive plant species. Binding of tentoxin to a high affinity (Ki ≈ 10 nM) site on the chloroplast F1 (CF1) strongly inhibits catalytic function, whereas binding to a second, lower affinity site (Kd > 10 μM) leads to restoration and even stimulation of catalytic activity. Sensitivity to tentoxin has been shown to be due, in part, to the nature of the amino acid residue at position 83 on the catalytic β subunit of CF1. An aspartate in this position is required, but is not sufficient, for tentoxin inhibition. By comparison with the solved structure of mitochondrial F1 [Abrahams, J. P., Leslie, A. G. W., Lutter, R., and Walker, J. E. (1994) Nature 370, 621-628], Asp83 is probably located at an interface between α and β subunits on CF1 where residues on the α subunit could also participate in tentoxin binding. A hybrid core F1 enzyme assembled with β and γ subunits of the tentoxin-sensitive spinach CF1, and an α subunit of the tentoxin-insensitive photosynthetic bacterium Rhodospirillum rubrum F1 (RrF1), was stimulated but not inhibited by tentoxin [Tucker, W. C., Du, Z., Gromet-Elhanan, Z. and Richter, M. L. (2001) Eur. J. Biochem. 268, 2179-2186]. In this study, chimeric α subunits were prepared by introducing short segments of the spinach CF1 α subunit from a poorly conserved region which is immediately adjacent to β-Asp83 in the crystal structure, into equivalent positions in the RrF1 α subunit using oligonucleotide-directed mutagenesis. Hybrid enzymes containing these chimeric α subunits had both the high affinity inhibitory tentoxin binding site and the lower affinity stimulatory site. Changing β-Asp83 to leucine resulted in loss of both inhibition and stimulation by tentoxin in the chimeras. The results indicate that tentoxin inhibition requires additional α residues that are not present on the RrF1 α subunit. A structural model of a putative inhibitory tentoxin binding pocket is presented.

UR - http://www.scopus.com/inward/record.url?scp=0035954403&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0035954403&partnerID=8YFLogxK

U2 - 10.1021/bi0105227

DO - 10.1021/bi0105227

M3 - Article

VL - 40

SP - 7542

EP - 7548

JO - Biochemistry

JF - Biochemistry

SN - 0006-2960

IS - 25

ER -