Inhibition of Pro-Apoptotic BAX by a Noncanonical Interaction Mechanism

Lauren A. Barclay; Thomas E. Wales; Thomas P. Garner; Franziska Wachter; Susan Lee; Rachel M. Guerra; Michelle L. Stewart; Craig R. Braun; Gregory H. Bird; Evripidis Gavathiotis; John R. Engen; Loren D. Walensky

doi:10.1016/j.molcel.2015.01.014

Inhibition of Pro-Apoptotic BAX by a Noncanonical Interaction Mechanism

Lauren A. Barclay, Thomas E. Wales, Thomas P. Garner, Franziska Wachter, Susan Lee, Rachel M. Guerra, Michelle L. Stewart, Craig R. Braun, Gregory H. Bird, Evripidis Gavathiotis, John R. Engen, Loren D. Walensky

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

108 Scopus citations

Abstract

BCL-2 is a negative regulator of apoptosis implicated in homeostatic and pathologic cell survival. The canonical anti-apoptotic mechanism involves entrapment of activated BAX by a groove on BCL-2, preventing BAX homo-oligomerization and mitochondrial membrane poration. The BCL-2 BH4 domain also confers anti-apoptotic functionality, but the mechanism is unknown. We find that a synthetic α-helical BH4 domain binds to BAX with nanomolar affinity and independently inhibits the conformational activation of BAX. Hydrogen-deuterium exchange mass spectrometry demonstrated that the N-terminal conformational changes in BAX induced by a triggering BIM BH3 helix were suppressed by the BCL-2 BH4 helix. Structural analyses localized the BH4 interaction site to a groove formed by residues of α1, α1-α2 loop, and α2-α3 and α5-α6 hairpins on the BAX surface. These data reveal a previously unappreciated binding site for targeted inhibition of BAX and suggest that the BCL-2 BH4 domain may participate in apoptosis blockade by a noncanonical interaction mechanism.

Original language	English (US)
Pages (from-to)	873-886
Number of pages	14
Journal	Molecular Cell
Volume	57
Issue number	5
DOIs	https://doi.org/10.1016/j.molcel.2015.01.014
State	Published - Mar 5 2015
Externally published	Yes

ASJC Scopus subject areas

Molecular Biology
Cell Biology

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10.1016/j.molcel.2015.01.014

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@article{3445d5390ce742f99fb9896f696b0098,

title = "Inhibition of Pro-Apoptotic BAX by a Noncanonical Interaction Mechanism",

abstract = "BCL-2 is a negative regulator of apoptosis implicated in homeostatic and pathologic cell survival. The canonical anti-apoptotic mechanism involves entrapment of activated BAX by a groove on BCL-2, preventing BAX homo-oligomerization and mitochondrial membrane poration. The BCL-2 BH4 domain also confers anti-apoptotic functionality, but the mechanism is unknown. We find that a synthetic α-helical BH4 domain binds to BAX with nanomolar affinity and independently inhibits the conformational activation of BAX. Hydrogen-deuterium exchange mass spectrometry demonstrated that the N-terminal conformational changes in BAX induced by a triggering BIM BH3 helix were suppressed by the BCL-2 BH4 helix. Structural analyses localized the BH4 interaction site to a groove formed by residues of α1, α1-α2 loop, and α2-α3 and α5-α6 hairpins on the BAX surface. These data reveal a previously unappreciated binding site for targeted inhibition of BAX and suggest that the BCL-2 BH4 domain may participate in apoptosis blockade by a noncanonical interaction mechanism.",

author = "Barclay, {Lauren A.} and Wales, {Thomas E.} and Garner, {Thomas P.} and Franziska Wachter and Susan Lee and Guerra, {Rachel M.} and Stewart, {Michelle L.} and Braun, {Craig R.} and Bird, {Gregory H.} and Evripidis Gavathiotis and Engen, {John R.} and Walensky, {Loren D.}",

note = "Funding Information: We thank E. Smith for editorial and graphics assistance, and S. Cahill for NMR technical support (Albert Einstein College of Medicine Structural NMR Resource). This work was supported by NIH grants 5R01CA050239 and 5R01GM090299 to L.D.W., NIH grants R01GM086507 and R01GM101135 to J.R.E., an NSF Graduate Research Fellowship to L.A.B., additional support to L.A.B. from T32GM007306, and NIH grant R00HL095929, a Kimmel Scholar Award, and a Sinsheimer Scholar Award to E.G. Additionally, E.G is a member of the New York Structural Biology Center (NYSBC), and the data collected at NYSBC was made possible by a grant from NYSTAR. L.D.W. is a scientific advisory board member and consultant for Aileron Therapeutics. J.R.E. is a consultant for the Waters Corporation. Publisher Copyright: {\textcopyright} 2015 Elsevier Inc.",

year = "2015",

month = mar,

day = "5",

doi = "10.1016/j.molcel.2015.01.014",

language = "English (US)",

volume = "57",

pages = "873--886",

journal = "Molecular Cell",

issn = "1097-2765",

publisher = "Cell Press",

number = "5",

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TY - JOUR

T1 - Inhibition of Pro-Apoptotic BAX by a Noncanonical Interaction Mechanism

AU - Barclay, Lauren A.

AU - Wales, Thomas E.

AU - Garner, Thomas P.

AU - Wachter, Franziska

AU - Lee, Susan

AU - Guerra, Rachel M.

AU - Stewart, Michelle L.

AU - Braun, Craig R.

AU - Bird, Gregory H.

AU - Gavathiotis, Evripidis

AU - Engen, John R.

AU - Walensky, Loren D.

N1 - Funding Information: We thank E. Smith for editorial and graphics assistance, and S. Cahill for NMR technical support (Albert Einstein College of Medicine Structural NMR Resource). This work was supported by NIH grants 5R01CA050239 and 5R01GM090299 to L.D.W., NIH grants R01GM086507 and R01GM101135 to J.R.E., an NSF Graduate Research Fellowship to L.A.B., additional support to L.A.B. from T32GM007306, and NIH grant R00HL095929, a Kimmel Scholar Award, and a Sinsheimer Scholar Award to E.G. Additionally, E.G is a member of the New York Structural Biology Center (NYSBC), and the data collected at NYSBC was made possible by a grant from NYSTAR. L.D.W. is a scientific advisory board member and consultant for Aileron Therapeutics. J.R.E. is a consultant for the Waters Corporation. Publisher Copyright: © 2015 Elsevier Inc.

PY - 2015/3/5

Y1 - 2015/3/5

N2 - BCL-2 is a negative regulator of apoptosis implicated in homeostatic and pathologic cell survival. The canonical anti-apoptotic mechanism involves entrapment of activated BAX by a groove on BCL-2, preventing BAX homo-oligomerization and mitochondrial membrane poration. The BCL-2 BH4 domain also confers anti-apoptotic functionality, but the mechanism is unknown. We find that a synthetic α-helical BH4 domain binds to BAX with nanomolar affinity and independently inhibits the conformational activation of BAX. Hydrogen-deuterium exchange mass spectrometry demonstrated that the N-terminal conformational changes in BAX induced by a triggering BIM BH3 helix were suppressed by the BCL-2 BH4 helix. Structural analyses localized the BH4 interaction site to a groove formed by residues of α1, α1-α2 loop, and α2-α3 and α5-α6 hairpins on the BAX surface. These data reveal a previously unappreciated binding site for targeted inhibition of BAX and suggest that the BCL-2 BH4 domain may participate in apoptosis blockade by a noncanonical interaction mechanism.

AB - BCL-2 is a negative regulator of apoptosis implicated in homeostatic and pathologic cell survival. The canonical anti-apoptotic mechanism involves entrapment of activated BAX by a groove on BCL-2, preventing BAX homo-oligomerization and mitochondrial membrane poration. The BCL-2 BH4 domain also confers anti-apoptotic functionality, but the mechanism is unknown. We find that a synthetic α-helical BH4 domain binds to BAX with nanomolar affinity and independently inhibits the conformational activation of BAX. Hydrogen-deuterium exchange mass spectrometry demonstrated that the N-terminal conformational changes in BAX induced by a triggering BIM BH3 helix were suppressed by the BCL-2 BH4 helix. Structural analyses localized the BH4 interaction site to a groove formed by residues of α1, α1-α2 loop, and α2-α3 and α5-α6 hairpins on the BAX surface. These data reveal a previously unappreciated binding site for targeted inhibition of BAX and suggest that the BCL-2 BH4 domain may participate in apoptosis blockade by a noncanonical interaction mechanism.

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U2 - 10.1016/j.molcel.2015.01.014

DO - 10.1016/j.molcel.2015.01.014

M3 - Article

C2 - 25684204

AN - SCOPUS:84924044826

SN - 1097-2765

VL - 57

SP - 873

EP - 886

JO - Molecular Cell

JF - Molecular Cell

IS - 5

ER -

Inhibition of Pro-Apoptotic BAX by a Noncanonical Interaction Mechanism

Abstract

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