A New Scaffold of an Old Protein Fold Ensures Binding to the Bisintercalator Thiocoraline

Tapan Biswas; Olga E. Zolova; Felipe Lombó; Fernando de la Calle; Jose A. Salas; Oleg V. Tsodikov; Sylvie Garneau-Tsodikova

doi:10.1016/j.jmb.2010.01.053

A New Scaffold of an Old Protein Fold Ensures Binding to the Bisintercalator Thiocoraline

Tapan Biswas, Olga E. Zolova, Felipe Lombó, Fernando de la Calle, Jose A. Salas, Oleg V. Tsodikov, Sylvie Garneau-Tsodikova

Research output: Contribution to journal › Article › peer-review

21 Scopus citations

Abstract

Thiocoraline is a thiodepsipeptide with potent antitumor activity. TioX, a protein with an unidentified function, is encoded by a gene of the thiocoraline biosynthetic gene cluster. The crystal structure of the full-length TioX protein at 2.15 Å resolution reveals that TioX protomer shares an ancient βαβββ fold motif with glyoxalase I and bleomycin resistance protein families, despite a very low sequence homology. Intriguingly, four TioX monomers form a unique 2-fold symmetric tetrameric assembly that is stabilized by four intermolecular disulfide bonds formed cyclically between Cys60 and Cys66 of adjacent monomers. The arrangement of two of the four monomers in the TioX tetramer is analogous to that in dimeric bleomycin resistance proteins. This analogy indicates that this novel higher-order structural scaffold of TioX may have evolved to bind thiocoraline. Our equilibrium titration studies demonstrate the binding of a thiocoraline chromophore analog, quinaldic acid, to TioX, thereby substantiating this model. Furthermore, a strain of Streptomyces albus containing an exogenous thiocoraline gene cluster devoid of functional tioX maintains thiocoraline production, albeit with a lower yield. Taken together, these observations rule out a direct enzymatic function of TioX and suggest that TioX is involved in thiocoraline resistance or secretion.

Original language	English
Pages (from-to)	495-507
Number of pages	13
Journal	Journal of Molecular Biology
Volume	397
Issue number	2
DOIs	https://doi.org/10.1016/j.jmb.2010.01.053
State	Published - Mar 26 2010

Bibliographical note

Funding Information:
This work was supported by start-up funds from the Life Sciences Institute (to S.G.-T.) and the College of Pharmacy (to S.G.-T. and O.V.T.) at the University of Michigan and by the Vahlteich Research Award Fund, College of Pharmacy at the University of Michigan (to S.G.-T.). Research performed at JAS laboratory was funded by Red Temática de Investigación Cooperativa de Centros de Cáncer (ISCIII-RETIC RD06/0020/0026; Ministry of Health, Spain). We thank Jessica M. Pero for the cloning of the His-tagged wild-type TioX and for the preparation of its SeMet-substituted variant. We thank Dr. Spencer Anderson and the staff of sectors LS-CAT and GM/CA-CAT at the Advanced Photon Source at the Argonne National Laboratory for assistance with the X-ray diffraction data collection.

Funding

This work was supported by start-up funds from the Life Sciences Institute (to S.G.-T.) and the College of Pharmacy (to S.G.-T. and O.V.T.) at the University of Michigan and by the Vahlteich Research Award Fund, College of Pharmacy at the University of Michigan (to S.G.-T.). Research performed at JAS laboratory was funded by Red Temática de Investigación Cooperativa de Centros de Cáncer (ISCIII-RETIC RD06/0020/0026; Ministry of Health, Spain). We thank Jessica M. Pero for the cloning of the His-tagged wild-type TioX and for the preparation of its SeMet-substituted variant. We thank Dr. Spencer Anderson and the staff of sectors LS-CAT and GM/CA-CAT at the Advanced Photon Source at the Argonne National Laboratory for assistance with the X-ray diffraction data collection.

Funders	Funder number
Red Temática de Investigación Cooperativa de Centros de Cáncer	ISCIII-RETIC RD06/0020/0026
University of Michigan Hospital
Institute of Life Sciences India
Ministerio de Sanidad, Servicios Sociales e Igualdad

Keywords

bacterial resistance
bisintercalator
hydrophobic pocket
thiocoraline
tryptophan fluorescence

ASJC Scopus subject areas

Structural Biology
Molecular Biology

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10.1016/j.jmb.2010.01.053

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title = "A New Scaffold of an Old Protein Fold Ensures Binding to the Bisintercalator Thiocoraline",

abstract = "Thiocoraline is a thiodepsipeptide with potent antitumor activity. TioX, a protein with an unidentified function, is encoded by a gene of the thiocoraline biosynthetic gene cluster. The crystal structure of the full-length TioX protein at 2.15 {\AA} resolution reveals that TioX protomer shares an ancient βαβββ fold motif with glyoxalase I and bleomycin resistance protein families, despite a very low sequence homology. Intriguingly, four TioX monomers form a unique 2-fold symmetric tetrameric assembly that is stabilized by four intermolecular disulfide bonds formed cyclically between Cys60 and Cys66 of adjacent monomers. The arrangement of two of the four monomers in the TioX tetramer is analogous to that in dimeric bleomycin resistance proteins. This analogy indicates that this novel higher-order structural scaffold of TioX may have evolved to bind thiocoraline. Our equilibrium titration studies demonstrate the binding of a thiocoraline chromophore analog, quinaldic acid, to TioX, thereby substantiating this model. Furthermore, a strain of Streptomyces albus containing an exogenous thiocoraline gene cluster devoid of functional tioX maintains thiocoraline production, albeit with a lower yield. Taken together, these observations rule out a direct enzymatic function of TioX and suggest that TioX is involved in thiocoraline resistance or secretion.",

keywords = "bacterial resistance, bisintercalator, hydrophobic pocket, thiocoraline, tryptophan fluorescence",

author = "Tapan Biswas and Zolova, {Olga E.} and Felipe Lomb{\'o} and {de la Calle}, Fernando and Salas, {Jose A.} and Tsodikov, {Oleg V.} and Sylvie Garneau-Tsodikova",

note = "Funding Information: This work was supported by start-up funds from the Life Sciences Institute (to S.G.-T.) and the College of Pharmacy (to S.G.-T. and O.V.T.) at the University of Michigan and by the Vahlteich Research Award Fund, College of Pharmacy at the University of Michigan (to S.G.-T.). Research performed at JAS laboratory was funded by Red Tem{\'a}tica de Investigaci{\'o}n Cooperativa de Centros de C{\'a}ncer (ISCIII-RETIC RD06/0020/0026; Ministry of Health, Spain). We thank Jessica M. Pero for the cloning of the His-tagged wild-type TioX and for the preparation of its SeMet-substituted variant. We thank Dr. Spencer Anderson and the staff of sectors LS-CAT and GM/CA-CAT at the Advanced Photon Source at the Argonne National Laboratory for assistance with the X-ray diffraction data collection. ",

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AU - Biswas, Tapan

AU - Zolova, Olga E.

AU - Lombó, Felipe

AU - de la Calle, Fernando

AU - Salas, Jose A.

AU - Tsodikov, Oleg V.

AU - Garneau-Tsodikova, Sylvie

N1 - Funding Information: This work was supported by start-up funds from the Life Sciences Institute (to S.G.-T.) and the College of Pharmacy (to S.G.-T. and O.V.T.) at the University of Michigan and by the Vahlteich Research Award Fund, College of Pharmacy at the University of Michigan (to S.G.-T.). Research performed at JAS laboratory was funded by Red Temática de Investigación Cooperativa de Centros de Cáncer (ISCIII-RETIC RD06/0020/0026; Ministry of Health, Spain). We thank Jessica M. Pero for the cloning of the His-tagged wild-type TioX and for the preparation of its SeMet-substituted variant. We thank Dr. Spencer Anderson and the staff of sectors LS-CAT and GM/CA-CAT at the Advanced Photon Source at the Argonne National Laboratory for assistance with the X-ray diffraction data collection.

PY - 2010/3/26

Y1 - 2010/3/26

N2 - Thiocoraline is a thiodepsipeptide with potent antitumor activity. TioX, a protein with an unidentified function, is encoded by a gene of the thiocoraline biosynthetic gene cluster. The crystal structure of the full-length TioX protein at 2.15 Å resolution reveals that TioX protomer shares an ancient βαβββ fold motif with glyoxalase I and bleomycin resistance protein families, despite a very low sequence homology. Intriguingly, four TioX monomers form a unique 2-fold symmetric tetrameric assembly that is stabilized by four intermolecular disulfide bonds formed cyclically between Cys60 and Cys66 of adjacent monomers. The arrangement of two of the four monomers in the TioX tetramer is analogous to that in dimeric bleomycin resistance proteins. This analogy indicates that this novel higher-order structural scaffold of TioX may have evolved to bind thiocoraline. Our equilibrium titration studies demonstrate the binding of a thiocoraline chromophore analog, quinaldic acid, to TioX, thereby substantiating this model. Furthermore, a strain of Streptomyces albus containing an exogenous thiocoraline gene cluster devoid of functional tioX maintains thiocoraline production, albeit with a lower yield. Taken together, these observations rule out a direct enzymatic function of TioX and suggest that TioX is involved in thiocoraline resistance or secretion.

AB - Thiocoraline is a thiodepsipeptide with potent antitumor activity. TioX, a protein with an unidentified function, is encoded by a gene of the thiocoraline biosynthetic gene cluster. The crystal structure of the full-length TioX protein at 2.15 Å resolution reveals that TioX protomer shares an ancient βαβββ fold motif with glyoxalase I and bleomycin resistance protein families, despite a very low sequence homology. Intriguingly, four TioX monomers form a unique 2-fold symmetric tetrameric assembly that is stabilized by four intermolecular disulfide bonds formed cyclically between Cys60 and Cys66 of adjacent monomers. The arrangement of two of the four monomers in the TioX tetramer is analogous to that in dimeric bleomycin resistance proteins. This analogy indicates that this novel higher-order structural scaffold of TioX may have evolved to bind thiocoraline. Our equilibrium titration studies demonstrate the binding of a thiocoraline chromophore analog, quinaldic acid, to TioX, thereby substantiating this model. Furthermore, a strain of Streptomyces albus containing an exogenous thiocoraline gene cluster devoid of functional tioX maintains thiocoraline production, albeit with a lower yield. Taken together, these observations rule out a direct enzymatic function of TioX and suggest that TioX is involved in thiocoraline resistance or secretion.

KW - bacterial resistance

KW - bisintercalator

KW - hydrophobic pocket

KW - thiocoraline

KW - tryptophan fluorescence

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A New Scaffold of an Old Protein Fold Ensures Binding to the Bisintercalator Thiocoraline

Abstract

Bibliographical note

Funding

Keywords

ASJC Scopus subject areas

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