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Transcript of The role of H293 in Protein Arginine Methyltransferases 1 (PRMT1) Brittany Boykin Auburn University...
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The role of H293 in Protein Arginine Methyltransferases 1 (PRMT1)
Brittany Boykin Auburn University
Department of Chemistry and Biochemistry Computational Seminar
National Organization for the Professional Organization of Black Chemist and Chemical Engineers 2015 Annual Conference
Orlando, Florida24, September 2015
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OutlineBackground
Overall Goal
• Future Work• Acknowledgments • Q/A
Current Study
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Post-Translational Modification (PTMs)
• Remarkably, there are 200+ types of PTM’s that include kinases, phosphatases, transferases
Walsh C. (2006) Posttranslational Modification of Proteins: Expanding Natures Inventory. Englewood, Colo.: Roberts and Co. Publishers. Xxi, 490 p. p.
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Arginine Methylation
FASEB J. 10, 471-480 (1996)
• Methylation reactions involve methyl group transfers (AdoMet is the methyl donor and this reaction displays a Sn2 type geometry)
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Protein Arginine Methyltransferases (PRMTs)
Bedford, M. T., and Clarke, S. G. (2009) Protein Arginine Methylation in mammals: Who, What, and why. Mol Cell 33, 1-13
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Physiological roles of PRMTs
Transcriptional Activity
RNA splicing and transport DNA repair
Chromatin Remodeling
Signal Transduction
Neuronal Cell Differentiation
Virus-host Biochemistry
Journal of Biological Chemistry Vol. 289, NO. 13, pp. 9320-9327, March 28, (2014)
• PRMTs regulate proteins in cell processes and human diseases
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Target Diseases
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PRMT Human Isoforms
Cell. Mol. Life Sci. 2009 66:2109.
PRMT1
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Human Diseases: PRMT1’s Target
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Structure of PRMT1
Structure 2003 11: 509.
Dimerization - essential for SAM binding and enzymatic activity
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Active Site of PRMT1
Biochemistry 2011, 50, 3332-3345J. Bio. Chem. Vol. 289, NO. 13, pp. 9320-9327 2014
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Overall Goal Product Specificity
Org. Biomol. Chem., 2015, 13, 549-560
Control Sterics & Nucleophilicity
• What use of the product depends on the local conditions?
• We want to gain more insight by dissecting the active site of PRMT1 and identifying the significance of specific
residues in regards to the substrate
• What orientation does the substrate display with the
H293S mutation?
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Importance of H293
Salt-Bridge
Biochemistry 2011 April 26; 50(16): 3332-3345
• This short bond plays a critical role in forming the two-helix boundary that impact cofactor and peptide binding
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Alternative Mechanism
NNHNN
N
OO
H3C
S
H
SAM
His293
Glu144
HH
H H
Arg-substrate
O
O
Glu153
NNHNN
N
OOS
H
SAM
His293
Glu144
H
H
H3C H
dicationintermediate
O
O
Glu153
H
NNHNN
N
OOS
H
SAM
His293
Glu144
H
H3C H
MMA-productO
O
Glu153
H
H
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Current Research Objective Objective • Product Specificity of H293: Mutant H293S;
• How does the protein environment influence product specificity using aMD simulations and QM/MM calculations
• Analyzed systems:• H293S-Arg• H293S-MMA-ADMA• H293S-MMA-SDMA
• With the Ser in place of His:• What orientation does the substrate display• Distribution of the His vs Ser
Question:• Does H293 have more affect on the active site than
proposed?
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Assisted Model Building With Energy Refinement (AMBER)
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Molecular Dynamics (MD)
F = ma
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(QM) | (MM)
• Quantum Mechanical (QM) Electronic Processes Bonding Breaking/formation DFT/ Ab Initio Primary subsystem (PS)
• Molecular Mechanical (MM) Force-field based method Computationally Efficient Secondary subsystem (SS)
• Combined QM/MM Chemical Reaction in macromolecules
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PRMT1 RMSF Analysis
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Preferred Methylation
13.512.010.59.07.56.04.5
1.2
1.0
0.8
0.6
0.4
0.2
0.0
d+0.5*cos(angle) radians
Den
sity
H293S-MMA-Nn2-P1
13.512.010.59.07.56.04.5
1.2
1.0
0.8
0.6
0.4
0.2
0.0
d+0.5*cos(angle) radians_1
Den
sity
H293S-MMA-Nn2-P2
13.0511.6010.158.707.255.804.35
1.2
1.0
0.8
0.6
0.4
0.2
0.0
d+0.5*cos(angle) radians
Den
sity
H293S-MMA-Nn1-P1
13.0511.6010.158.707.255.804.35
1.2
1.0
0.8
0.6
0.4
0.2
0.0
d+0.5*cos(angle) radians
Den
sity
H293S-MMA-Nn1-P2
SDMA
ADMA
ADMASDMA
H293S-MMANη2
H293S-MMANη1
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Future Work
• To continue the QM/MM simulations for all mutant complexes: H293S-Arg, H293S-MMA-proADMA, and H293S-MMA-proSDMA
• Continue to compare to double mutant (H293S-M48F) and WT-enzyme
• To compare to experimental results
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AcknowledgementsoAlabama Super Computer Center oHuntsville Alabama
oCollaborators: Dr. Joan Hevel (Utah State University)oOrlando AcevedooMy Lab members oSymon GathiakaoNicole IppolitooRobel GhebreaboBrian Doherty
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