Solution Structure of the Integral Human Membrane Protein VDAC-1

in Detergent Micelles

Presented by Lisa Nguyen

Sebastian Hiller1 Robert G Garces1 Thomas J Malia1dagger Vladislav Y Orekhov13Marco Colombini2 Gerhard Wagner1Dagger

IntroductionMitochondriabullProduction of cellular energy via ATP synthase and ETCbullOther Metabolic Activities

ndash Regulation of the membrane potential

ndash Apoptosisndash Cellular proliferation

and differentiationndash Heme Synthesis

Voltage-dependent anion channel (VDAC)

bull Integral membrane protein

bull Mitochondrial porin bull Three isoforms

VDAC-1 VDAC-2 VDAC-3

bull Diffusion of small hydrophilic molecules

bull Conserved across eukaryotes (30 similarity between yeast and human)

Structure and Functions of VDAC-1

bull 19-stranded β barrelbull Short α helixbull A parallel β-strand pairing bull N- and C- terminus are on opposite sidebull Open conformation at low membrane potential ~10mVbull Closed conformation at ~30mVbull VDAC mediates traffic of small moleculesbull Involved in apoptotic pathways (via interaction with Bcl-2 family )

Ujwal R et al PNAS 200810517742-17747

IntroductionMitochondriabullProduction of cellular energy via ATP synthase and ETCbullOther Metabolic Activities

ndash Regulation of the membrane potential

ndash Apoptosisndash Cellular proliferation

and differentiationndash Heme Synthesis

Voltage-dependent anion channel (VDAC)

bull Integral membrane protein

bull Mitochondrial porin bull Three isoforms

VDAC-1 VDAC-2 VDAC-3

bull Diffusion of small hydrophilic molecules

bull Conserved across eukaryotes (30 similarity between yeast and human)

Structure and Functions of VDAC-1

bull 19-stranded β barrelbull Short α helixbull A parallel β-strand pairing bull N- and C- terminus are on opposite sidebull Open conformation at low membrane potential ~10mVbull Closed conformation at ~30mVbull VDAC mediates traffic of small moleculesbull Involved in apoptotic pathways (via interaction with Bcl-2 family )

Ujwal R et al PNAS 200810517742-17747

Voltage-dependent anion channel (VDAC)

bull Integral membrane protein

bull Mitochondrial porin bull Three isoforms

VDAC-1 VDAC-2 VDAC-3

bull Diffusion of small hydrophilic molecules

bull Conserved across eukaryotes (30 similarity between yeast and human)

Structure and Functions of VDAC-1

bull 19-stranded β barrelbull Short α helixbull A parallel β-strand pairing bull N- and C- terminus are on opposite sidebull Open conformation at low membrane potential ~10mVbull Closed conformation at ~30mVbull VDAC mediates traffic of small moleculesbull Involved in apoptotic pathways (via interaction with Bcl-2 family )

Ujwal R et al PNAS 200810517742-17747

Structure and Functions of VDAC-1

bull 19-stranded β barrelbull Short α helixbull A parallel β-strand pairing bull N- and C- terminus are on opposite sidebull Open conformation at low membrane potential ~10mVbull Closed conformation at ~30mVbull VDAC mediates traffic of small moleculesbull Involved in apoptotic pathways (via interaction with Bcl-2 family )

Ujwal R et al PNAS 200810517742-17747

Bcl-2 Family of Proteinbull Involved in apoptosis (Bcl-xL Bax and Bak)bull Bcl-xL

ndash Antiapoptosisndash Binding to VDAC -1 opens the channelndash Inhibit the release of apoptogenic proteins (cytochrome c)

Defects in Bcl-2 lead tobullCancer bullCardiovascular diseasesbullNeurological diseases

Studies Overview bull 3D solution structure of VDAC-1

reconstituted in detergent micelles LDAO in solution

bull Nuclear magnetic resonance (NMR)ndash TROSY (Transverse Relaxation-

Optimized Spectroscopy) ndash NOESY (Nuclear Overhauser

Effect Spectroscopy)bull NMR measurements revealed

the binding sites of VDAC-1 for the Bcl-2 protein Bcl-xL for reduced βndashnicotinamide adenine dinucleotide (β-NADH) and for cholesterol

Architecture of VDAC-1

S Hiller et al Science 20083211206-1210

Published by AAAS

NMR solution structure of VDAC-1 in LDAO micelles

S Hiller et al Science 20083211206-1210

Published by AAAS

Proposed model of the transition from open to closed state

Ujwal R et al PNAS 200810517742-17747

copy2008 by National Academy of Sciences

Hydrophobic surface of VDAC-1

S Hiller et al Science 20083211206-1210

Published by AAAS

Limitation of the experiment

bull Cannot tell which opening of the channel faces into the cytosol and which faces into mitochondria

bull The exact structure and localization of the N-terminal segment cannot be determined due to unassigned regions of the N-terminus

bull The data excludes formation of stable oligomers in the LDAO micelles

Cholesterol β-NADH and Bcl-xL

bull In the presence of cholesterol recombinant VDAC-1 can form voltage-gated channels in phospholipid bilayers similar to those of the native protein

bull ATP and β-NAD do not interact with a specific site of the VDAC-1

bull β -NADH interacts with VDAC-1 at strands 17 and 18bull β -NADH favors the closure of VDAC which

limits metabolite flux across the outer membrane and inhibits mitochondrial function

bull Bcl-xL binds to the VDAC-1 at strands 17 and 18ndash Bcl-xL

provides protection against apoptosis through interaction with the VDAC-1

Interactions of VDAC-1

S Hiller et al Science 20083211206-1210

Published by AAAS

Conclusion

bull VDAC1 was shown to adopt a β-barrel architecture comprising 19 β-strands and an α-helix located within the pore The α-helix of the N-terminal segment is oriented against the interior wall causing a partial narrowing at the center of the pore

bull Studies of VDAC-1 structures suggest that the hydrophilic N-terminus of the protein is nestled within the pore

bull N-terminal α -helix acts as a voltage gating

bull β-NADH and Bcl-xL interacts with VDAC-1 in its open conformation

Table S1 NMR structure statistics

NOE distance restraints All HN-HN HN-Meth Meth-Meth

Intraresidual 69 0 69 0

Sequential 199 129 69 1

Medium range (2 le i-j le 4) 72 28 34 10

Long range (i-jgt4) 272 131 85 56

Total 612 288 257 67

Hydrogen bond restraints 139

Dihedral angle restraints (φ and ψ) 2 x 158

Ramachandran plot

Most favored region 771

Additionally allowed region 216

generously allowed region 08

Disallowed region 04

Deviations from idealized geometry

Bond lengths (Aring) 0005

Bond angles (deg) 07

Average pairwise rmsd (backbone Aring)

All residues except 1ndash25 36

All residues except 1ndash25 and loopsa 29

• Solution Structure of the Integral Human Membrane Protein VDAC-1 in Detergent Micelles
• Introduction
• Voltage-dependent anion channel (VDAC)
• Structure and Functions of VDAC-1
• PowerPoint Presentation
• Slide 6
• Bcl-2 Family of Protein
• Studies Overview
• Slide 9
• Slide 10
• Slide 11
• Slide 12
• Limitation of the experiment
• Cholesterol β-NADH and Bcl-xL
• Slide 15
• Conclusion
• Slide 17

Studies Overview bull 3D solution structure of VDAC-1

reconstituted in detergent micelles LDAO in solution

bull Nuclear magnetic resonance (NMR)ndash TROSY (Transverse Relaxation-

Optimized Spectroscopy) ndash NOESY (Nuclear Overhauser

Effect Spectroscopy)bull NMR measurements revealed

the binding sites of VDAC-1 for the Bcl-2 protein Bcl-xL for reduced βndashnicotinamide adenine dinucleotide (β-NADH) and for cholesterol

Architecture of VDAC-1

S Hiller et al Science 20083211206-1210

Published by AAAS

NMR solution structure of VDAC-1 in LDAO micelles

S Hiller et al Science 20083211206-1210

Published by AAAS

Proposed model of the transition from open to closed state

Ujwal R et al PNAS 200810517742-17747

copy2008 by National Academy of Sciences

Hydrophobic surface of VDAC-1

S Hiller et al Science 20083211206-1210

Published by AAAS

Limitation of the experiment

bull Cannot tell which opening of the channel faces into the cytosol and which faces into mitochondria

bull The exact structure and localization of the N-terminal segment cannot be determined due to unassigned regions of the N-terminus

bull The data excludes formation of stable oligomers in the LDAO micelles

Cholesterol β-NADH and Bcl-xL

bull In the presence of cholesterol recombinant VDAC-1 can form voltage-gated channels in phospholipid bilayers similar to those of the native protein

bull ATP and β-NAD do not interact with a specific site of the VDAC-1

bull β -NADH interacts with VDAC-1 at strands 17 and 18bull β -NADH favors the closure of VDAC which

limits metabolite flux across the outer membrane and inhibits mitochondrial function

bull Bcl-xL binds to the VDAC-1 at strands 17 and 18ndash Bcl-xL

provides protection against apoptosis through interaction with the VDAC-1

Interactions of VDAC-1

S Hiller et al Science 20083211206-1210

Published by AAAS

Conclusion

bull VDAC1 was shown to adopt a β-barrel architecture comprising 19 β-strands and an α-helix located within the pore The α-helix of the N-terminal segment is oriented against the interior wall causing a partial narrowing at the center of the pore

bull Studies of VDAC-1 structures suggest that the hydrophilic N-terminus of the protein is nestled within the pore

bull N-terminal α -helix acts as a voltage gating

bull β-NADH and Bcl-xL interacts with VDAC-1 in its open conformation

Table S1 NMR structure statistics

NOE distance restraints All HN-HN HN-Meth Meth-Meth

Intraresidual 69 0 69 0

Sequential 199 129 69 1

Medium range (2 le i-j le 4) 72 28 34 10

Long range (i-jgt4) 272 131 85 56

Total 612 288 257 67

Hydrogen bond restraints 139

Dihedral angle restraints (φ and ψ) 2 x 158

Ramachandran plot

Most favored region 771

Additionally allowed region 216

generously allowed region 08

Disallowed region 04

Deviations from idealized geometry

Bond lengths (Aring) 0005

Bond angles (deg) 07

Average pairwise rmsd (backbone Aring)

All residues except 1ndash25 36

All residues except 1ndash25 and loopsa 29

• Solution Structure of the Integral Human Membrane Protein VDAC-1 in Detergent Micelles
• Introduction
• Voltage-dependent anion channel (VDAC)
• Structure and Functions of VDAC-1
• PowerPoint Presentation
• Slide 6
• Bcl-2 Family of Protein
• Studies Overview
• Slide 9
• Slide 10
• Slide 11
• Slide 12
• Limitation of the experiment
• Cholesterol β-NADH and Bcl-xL
• Slide 15
• Conclusion
• Slide 17

Architecture of VDAC-1

S Hiller et al Science 20083211206-1210

Published by AAAS

NMR solution structure of VDAC-1 in LDAO micelles

S Hiller et al Science 20083211206-1210

Published by AAAS

Proposed model of the transition from open to closed state

Ujwal R et al PNAS 200810517742-17747

copy2008 by National Academy of Sciences

Hydrophobic surface of VDAC-1

S Hiller et al Science 20083211206-1210

Published by AAAS

Limitation of the experiment

bull Cannot tell which opening of the channel faces into the cytosol and which faces into mitochondria

bull The exact structure and localization of the N-terminal segment cannot be determined due to unassigned regions of the N-terminus

bull The data excludes formation of stable oligomers in the LDAO micelles

Cholesterol β-NADH and Bcl-xL

bull In the presence of cholesterol recombinant VDAC-1 can form voltage-gated channels in phospholipid bilayers similar to those of the native protein

bull ATP and β-NAD do not interact with a specific site of the VDAC-1

bull β -NADH interacts with VDAC-1 at strands 17 and 18bull β -NADH favors the closure of VDAC which

limits metabolite flux across the outer membrane and inhibits mitochondrial function

bull Bcl-xL binds to the VDAC-1 at strands 17 and 18ndash Bcl-xL

provides protection against apoptosis through interaction with the VDAC-1

Interactions of VDAC-1

S Hiller et al Science 20083211206-1210

Published by AAAS

Conclusion

bull VDAC1 was shown to adopt a β-barrel architecture comprising 19 β-strands and an α-helix located within the pore The α-helix of the N-terminal segment is oriented against the interior wall causing a partial narrowing at the center of the pore

bull Studies of VDAC-1 structures suggest that the hydrophilic N-terminus of the protein is nestled within the pore

bull N-terminal α -helix acts as a voltage gating

bull β-NADH and Bcl-xL interacts with VDAC-1 in its open conformation

Table S1 NMR structure statistics

NOE distance restraints All HN-HN HN-Meth Meth-Meth

Intraresidual 69 0 69 0

Sequential 199 129 69 1

Medium range (2 le i-j le 4) 72 28 34 10

Long range (i-jgt4) 272 131 85 56

Total 612 288 257 67

Hydrogen bond restraints 139

Dihedral angle restraints (φ and ψ) 2 x 158

Ramachandran plot

Most favored region 771

Additionally allowed region 216

generously allowed region 08

Disallowed region 04

Deviations from idealized geometry

Bond lengths (Aring) 0005

Bond angles (deg) 07

Average pairwise rmsd (backbone Aring)

All residues except 1ndash25 36

All residues except 1ndash25 and loopsa 29

• Solution Structure of the Integral Human Membrane Protein VDAC-1 in Detergent Micelles
• Introduction
• Voltage-dependent anion channel (VDAC)
• Structure and Functions of VDAC-1
• PowerPoint Presentation
• Slide 6
• Bcl-2 Family of Protein
• Studies Overview
• Slide 9
• Slide 10
• Slide 11
• Slide 12
• Limitation of the experiment
• Cholesterol β-NADH and Bcl-xL
• Slide 15
• Conclusion
• Slide 17

NMR solution structure of VDAC-1 in LDAO micelles

S Hiller et al Science 20083211206-1210

Published by AAAS

Proposed model of the transition from open to closed state

Ujwal R et al PNAS 200810517742-17747

copy2008 by National Academy of Sciences

Hydrophobic surface of VDAC-1

S Hiller et al Science 20083211206-1210

Published by AAAS

Limitation of the experiment

bull Cannot tell which opening of the channel faces into the cytosol and which faces into mitochondria

bull The exact structure and localization of the N-terminal segment cannot be determined due to unassigned regions of the N-terminus

bull The data excludes formation of stable oligomers in the LDAO micelles

Cholesterol β-NADH and Bcl-xL

bull In the presence of cholesterol recombinant VDAC-1 can form voltage-gated channels in phospholipid bilayers similar to those of the native protein

bull ATP and β-NAD do not interact with a specific site of the VDAC-1

bull β -NADH interacts with VDAC-1 at strands 17 and 18bull β -NADH favors the closure of VDAC which

limits metabolite flux across the outer membrane and inhibits mitochondrial function

bull Bcl-xL binds to the VDAC-1 at strands 17 and 18ndash Bcl-xL

provides protection against apoptosis through interaction with the VDAC-1

Interactions of VDAC-1

S Hiller et al Science 20083211206-1210

Published by AAAS

Conclusion

bull VDAC1 was shown to adopt a β-barrel architecture comprising 19 β-strands and an α-helix located within the pore The α-helix of the N-terminal segment is oriented against the interior wall causing a partial narrowing at the center of the pore

bull Studies of VDAC-1 structures suggest that the hydrophilic N-terminus of the protein is nestled within the pore

bull N-terminal α -helix acts as a voltage gating

bull β-NADH and Bcl-xL interacts with VDAC-1 in its open conformation

Table S1 NMR structure statistics

NOE distance restraints All HN-HN HN-Meth Meth-Meth

Intraresidual 69 0 69 0

Sequential 199 129 69 1

Medium range (2 le i-j le 4) 72 28 34 10

Long range (i-jgt4) 272 131 85 56

Total 612 288 257 67

Hydrogen bond restraints 139

Dihedral angle restraints (φ and ψ) 2 x 158

Ramachandran plot

Most favored region 771

Additionally allowed region 216

generously allowed region 08

Disallowed region 04

Deviations from idealized geometry

Bond lengths (Aring) 0005

Bond angles (deg) 07

Average pairwise rmsd (backbone Aring)

All residues except 1ndash25 36

All residues except 1ndash25 and loopsa 29

• Solution Structure of the Integral Human Membrane Protein VDAC-1 in Detergent Micelles
• Introduction
• Voltage-dependent anion channel (VDAC)
• Structure and Functions of VDAC-1
• PowerPoint Presentation
• Slide 6
• Bcl-2 Family of Protein
• Studies Overview
• Slide 9
• Slide 10
• Slide 11
• Slide 12
• Limitation of the experiment
• Cholesterol β-NADH and Bcl-xL
• Slide 15
• Conclusion
• Slide 17

Proposed model of the transition from open to closed state

Ujwal R et al PNAS 200810517742-17747

copy2008 by National Academy of Sciences

Hydrophobic surface of VDAC-1

S Hiller et al Science 20083211206-1210

Published by AAAS

Limitation of the experiment

bull Cannot tell which opening of the channel faces into the cytosol and which faces into mitochondria

bull The exact structure and localization of the N-terminal segment cannot be determined due to unassigned regions of the N-terminus

bull The data excludes formation of stable oligomers in the LDAO micelles

Cholesterol β-NADH and Bcl-xL

bull In the presence of cholesterol recombinant VDAC-1 can form voltage-gated channels in phospholipid bilayers similar to those of the native protein

bull ATP and β-NAD do not interact with a specific site of the VDAC-1

bull β -NADH interacts with VDAC-1 at strands 17 and 18bull β -NADH favors the closure of VDAC which

limits metabolite flux across the outer membrane and inhibits mitochondrial function

bull Bcl-xL binds to the VDAC-1 at strands 17 and 18ndash Bcl-xL

provides protection against apoptosis through interaction with the VDAC-1

Interactions of VDAC-1

S Hiller et al Science 20083211206-1210

Published by AAAS

Conclusion

bull VDAC1 was shown to adopt a β-barrel architecture comprising 19 β-strands and an α-helix located within the pore The α-helix of the N-terminal segment is oriented against the interior wall causing a partial narrowing at the center of the pore

bull Studies of VDAC-1 structures suggest that the hydrophilic N-terminus of the protein is nestled within the pore

bull N-terminal α -helix acts as a voltage gating

bull β-NADH and Bcl-xL interacts with VDAC-1 in its open conformation

Table S1 NMR structure statistics

NOE distance restraints All HN-HN HN-Meth Meth-Meth

Intraresidual 69 0 69 0

Sequential 199 129 69 1

Medium range (2 le i-j le 4) 72 28 34 10

Long range (i-jgt4) 272 131 85 56

Total 612 288 257 67

Hydrogen bond restraints 139

Dihedral angle restraints (φ and ψ) 2 x 158

Ramachandran plot

Most favored region 771

Additionally allowed region 216

generously allowed region 08

Disallowed region 04

Deviations from idealized geometry

Bond lengths (Aring) 0005

Bond angles (deg) 07

Average pairwise rmsd (backbone Aring)

All residues except 1ndash25 36

All residues except 1ndash25 and loopsa 29

• Solution Structure of the Integral Human Membrane Protein VDAC-1 in Detergent Micelles
• Introduction
• Voltage-dependent anion channel (VDAC)
• Structure and Functions of VDAC-1
• PowerPoint Presentation
• Slide 6
• Bcl-2 Family of Protein
• Studies Overview
• Slide 9
• Slide 10
• Slide 11
• Slide 12
• Limitation of the experiment
• Cholesterol β-NADH and Bcl-xL
• Slide 15
• Conclusion
• Slide 17

Hydrophobic surface of VDAC-1

S Hiller et al Science 20083211206-1210

Published by AAAS

Limitation of the experiment

bull Cannot tell which opening of the channel faces into the cytosol and which faces into mitochondria

bull The exact structure and localization of the N-terminal segment cannot be determined due to unassigned regions of the N-terminus

bull The data excludes formation of stable oligomers in the LDAO micelles

Cholesterol β-NADH and Bcl-xL

bull In the presence of cholesterol recombinant VDAC-1 can form voltage-gated channels in phospholipid bilayers similar to those of the native protein

bull ATP and β-NAD do not interact with a specific site of the VDAC-1

bull β -NADH interacts with VDAC-1 at strands 17 and 18bull β -NADH favors the closure of VDAC which

limits metabolite flux across the outer membrane and inhibits mitochondrial function

bull Bcl-xL binds to the VDAC-1 at strands 17 and 18ndash Bcl-xL

provides protection against apoptosis through interaction with the VDAC-1

Interactions of VDAC-1

S Hiller et al Science 20083211206-1210

Published by AAAS

Conclusion

bull VDAC1 was shown to adopt a β-barrel architecture comprising 19 β-strands and an α-helix located within the pore The α-helix of the N-terminal segment is oriented against the interior wall causing a partial narrowing at the center of the pore

bull Studies of VDAC-1 structures suggest that the hydrophilic N-terminus of the protein is nestled within the pore

bull N-terminal α -helix acts as a voltage gating

bull β-NADH and Bcl-xL interacts with VDAC-1 in its open conformation

Table S1 NMR structure statistics

NOE distance restraints All HN-HN HN-Meth Meth-Meth

Intraresidual 69 0 69 0

Sequential 199 129 69 1

Medium range (2 le i-j le 4) 72 28 34 10

Long range (i-jgt4) 272 131 85 56

Total 612 288 257 67

Hydrogen bond restraints 139

Dihedral angle restraints (φ and ψ) 2 x 158

Ramachandran plot

Most favored region 771

Additionally allowed region 216

generously allowed region 08

Disallowed region 04

Deviations from idealized geometry

Bond lengths (Aring) 0005

Bond angles (deg) 07

Average pairwise rmsd (backbone Aring)

All residues except 1ndash25 36

All residues except 1ndash25 and loopsa 29

• Solution Structure of the Integral Human Membrane Protein VDAC-1 in Detergent Micelles
• Introduction
• Voltage-dependent anion channel (VDAC)
• Structure and Functions of VDAC-1
• PowerPoint Presentation
• Slide 6
• Bcl-2 Family of Protein
• Studies Overview
• Slide 9
• Slide 10
• Slide 11
• Slide 12
• Limitation of the experiment
• Cholesterol β-NADH and Bcl-xL
• Slide 15
• Conclusion
• Slide 17

Limitation of the experiment

bull Cannot tell which opening of the channel faces into the cytosol and which faces into mitochondria

bull The exact structure and localization of the N-terminal segment cannot be determined due to unassigned regions of the N-terminus

bull The data excludes formation of stable oligomers in the LDAO micelles

Cholesterol β-NADH and Bcl-xL

bull In the presence of cholesterol recombinant VDAC-1 can form voltage-gated channels in phospholipid bilayers similar to those of the native protein

bull ATP and β-NAD do not interact with a specific site of the VDAC-1

bull β -NADH interacts with VDAC-1 at strands 17 and 18bull β -NADH favors the closure of VDAC which

limits metabolite flux across the outer membrane and inhibits mitochondrial function

bull Bcl-xL binds to the VDAC-1 at strands 17 and 18ndash Bcl-xL

provides protection against apoptosis through interaction with the VDAC-1

Interactions of VDAC-1

S Hiller et al Science 20083211206-1210

Published by AAAS

Conclusion

bull VDAC1 was shown to adopt a β-barrel architecture comprising 19 β-strands and an α-helix located within the pore The α-helix of the N-terminal segment is oriented against the interior wall causing a partial narrowing at the center of the pore

bull Studies of VDAC-1 structures suggest that the hydrophilic N-terminus of the protein is nestled within the pore

bull N-terminal α -helix acts as a voltage gating

bull β-NADH and Bcl-xL interacts with VDAC-1 in its open conformation

Table S1 NMR structure statistics

NOE distance restraints All HN-HN HN-Meth Meth-Meth

Intraresidual 69 0 69 0

Sequential 199 129 69 1

Medium range (2 le i-j le 4) 72 28 34 10

Long range (i-jgt4) 272 131 85 56

Total 612 288 257 67

Hydrogen bond restraints 139

Dihedral angle restraints (φ and ψ) 2 x 158

Ramachandran plot

Most favored region 771

Additionally allowed region 216

generously allowed region 08

Disallowed region 04

Deviations from idealized geometry

Bond lengths (Aring) 0005

Bond angles (deg) 07

Average pairwise rmsd (backbone Aring)

All residues except 1ndash25 36

All residues except 1ndash25 and loopsa 29

• Solution Structure of the Integral Human Membrane Protein VDAC-1 in Detergent Micelles
• Introduction
• Voltage-dependent anion channel (VDAC)
• Structure and Functions of VDAC-1
• PowerPoint Presentation
• Slide 6
• Bcl-2 Family of Protein
• Studies Overview
• Slide 9
• Slide 10
• Slide 11
• Slide 12
• Limitation of the experiment
• Cholesterol β-NADH and Bcl-xL
• Slide 15
• Conclusion
• Slide 17

Cholesterol β-NADH and Bcl-xL

bull In the presence of cholesterol recombinant VDAC-1 can form voltage-gated channels in phospholipid bilayers similar to those of the native protein

bull ATP and β-NAD do not interact with a specific site of the VDAC-1

bull β -NADH interacts with VDAC-1 at strands 17 and 18bull β -NADH favors the closure of VDAC which

limits metabolite flux across the outer membrane and inhibits mitochondrial function

bull Bcl-xL binds to the VDAC-1 at strands 17 and 18ndash Bcl-xL

provides protection against apoptosis through interaction with the VDAC-1

Interactions of VDAC-1

S Hiller et al Science 20083211206-1210

Published by AAAS

Conclusion

bull VDAC1 was shown to adopt a β-barrel architecture comprising 19 β-strands and an α-helix located within the pore The α-helix of the N-terminal segment is oriented against the interior wall causing a partial narrowing at the center of the pore

bull Studies of VDAC-1 structures suggest that the hydrophilic N-terminus of the protein is nestled within the pore

bull N-terminal α -helix acts as a voltage gating

bull β-NADH and Bcl-xL interacts with VDAC-1 in its open conformation

Table S1 NMR structure statistics

NOE distance restraints All HN-HN HN-Meth Meth-Meth

Intraresidual 69 0 69 0

Sequential 199 129 69 1

Medium range (2 le i-j le 4) 72 28 34 10

Long range (i-jgt4) 272 131 85 56

Total 612 288 257 67

Hydrogen bond restraints 139

Dihedral angle restraints (φ and ψ) 2 x 158

Ramachandran plot

Most favored region 771

Additionally allowed region 216

generously allowed region 08

Disallowed region 04

Deviations from idealized geometry

Bond lengths (Aring) 0005

Bond angles (deg) 07

Average pairwise rmsd (backbone Aring)

All residues except 1ndash25 36

All residues except 1ndash25 and loopsa 29

• Solution Structure of the Integral Human Membrane Protein VDAC-1 in Detergent Micelles
• Introduction
• Voltage-dependent anion channel (VDAC)
• Structure and Functions of VDAC-1
• PowerPoint Presentation
• Slide 6
• Bcl-2 Family of Protein
• Studies Overview
• Slide 9
• Slide 10
• Slide 11
• Slide 12
• Limitation of the experiment
• Cholesterol β-NADH and Bcl-xL
• Slide 15
• Conclusion
• Slide 17

Interactions of VDAC-1

S Hiller et al Science 20083211206-1210

Published by AAAS

Conclusion

bull VDAC1 was shown to adopt a β-barrel architecture comprising 19 β-strands and an α-helix located within the pore The α-helix of the N-terminal segment is oriented against the interior wall causing a partial narrowing at the center of the pore

bull Studies of VDAC-1 structures suggest that the hydrophilic N-terminus of the protein is nestled within the pore

bull N-terminal α -helix acts as a voltage gating

bull β-NADH and Bcl-xL interacts with VDAC-1 in its open conformation

Table S1 NMR structure statistics

NOE distance restraints All HN-HN HN-Meth Meth-Meth

Intraresidual 69 0 69 0

Sequential 199 129 69 1

Medium range (2 le i-j le 4) 72 28 34 10

Long range (i-jgt4) 272 131 85 56

Total 612 288 257 67

Hydrogen bond restraints 139

Dihedral angle restraints (φ and ψ) 2 x 158

Ramachandran plot

Most favored region 771

Additionally allowed region 216

generously allowed region 08

Disallowed region 04

Deviations from idealized geometry

Bond lengths (Aring) 0005

Bond angles (deg) 07

Average pairwise rmsd (backbone Aring)

All residues except 1ndash25 36

All residues except 1ndash25 and loopsa 29

• Solution Structure of the Integral Human Membrane Protein VDAC-1 in Detergent Micelles
• Introduction
• Voltage-dependent anion channel (VDAC)
• Structure and Functions of VDAC-1
• PowerPoint Presentation
• Slide 6
• Bcl-2 Family of Protein
• Studies Overview
• Slide 9
• Slide 10
• Slide 11
• Slide 12
• Limitation of the experiment
• Cholesterol β-NADH and Bcl-xL
• Slide 15
• Conclusion
• Slide 17

Conclusion

bull VDAC1 was shown to adopt a β-barrel architecture comprising 19 β-strands and an α-helix located within the pore The α-helix of the N-terminal segment is oriented against the interior wall causing a partial narrowing at the center of the pore

bull Studies of VDAC-1 structures suggest that the hydrophilic N-terminus of the protein is nestled within the pore

bull N-terminal α -helix acts as a voltage gating

bull β-NADH and Bcl-xL interacts with VDAC-1 in its open conformation

Table S1 NMR structure statistics

NOE distance restraints All HN-HN HN-Meth Meth-Meth

Intraresidual 69 0 69 0

Sequential 199 129 69 1

Medium range (2 le i-j le 4) 72 28 34 10

Long range (i-jgt4) 272 131 85 56

Total 612 288 257 67

Hydrogen bond restraints 139

Dihedral angle restraints (φ and ψ) 2 x 158

Ramachandran plot

Most favored region 771

Additionally allowed region 216

generously allowed region 08

Disallowed region 04

Deviations from idealized geometry

Bond lengths (Aring) 0005

Bond angles (deg) 07

Average pairwise rmsd (backbone Aring)

All residues except 1ndash25 36

All residues except 1ndash25 and loopsa 29

• Solution Structure of the Integral Human Membrane Protein VDAC-1 in Detergent Micelles
• Introduction
• Voltage-dependent anion channel (VDAC)
• Structure and Functions of VDAC-1
• PowerPoint Presentation
• Slide 6
• Bcl-2 Family of Protein
• Studies Overview
• Slide 9
• Slide 10
• Slide 11
• Slide 12
• Limitation of the experiment
• Cholesterol β-NADH and Bcl-xL
• Slide 15
• Conclusion
• Slide 17

Table S1 NMR structure statistics

NOE distance restraints All HN-HN HN-Meth Meth-Meth

Intraresidual 69 0 69 0

Sequential 199 129 69 1

Medium range (2 le i-j le 4) 72 28 34 10

Long range (i-jgt4) 272 131 85 56

Total 612 288 257 67

Hydrogen bond restraints 139

Dihedral angle restraints (φ and ψ) 2 x 158

Ramachandran plot

Most favored region 771

Additionally allowed region 216

generously allowed region 08

Disallowed region 04

Deviations from idealized geometry

Bond lengths (Aring) 0005

Bond angles (deg) 07

Average pairwise rmsd (backbone Aring)

All residues except 1ndash25 36

All residues except 1ndash25 and loopsa 29

• Solution Structure of the Integral Human Membrane Protein VDAC-1 in Detergent Micelles
• Introduction
• Voltage-dependent anion channel (VDAC)
• Structure and Functions of VDAC-1
• PowerPoint Presentation
• Slide 6
• Bcl-2 Family of Protein
• Studies Overview
• Slide 9
• Slide 10
• Slide 11
• Slide 12
• Limitation of the experiment
• Cholesterol β-NADH and Bcl-xL
• Slide 15
• Conclusion
• Slide 17