Valyl-tRNA Synthetase Josh Jarodsky http://www.rcsb.org/pdb/education_discussion/molecule_of_the_month/images/aaRS.gif
Jan 20, 2016
Valyl-tRNA Synthetase
Josh Jarodskyhttp://www.rcsb.org/pdb/education_discussion/molecule_of_the_month/images/aaRS.gif
General Information
• Valyl-tRNA Synthetase = ValRS
• ValRS is an enzyme• E.C. 6.1.1.9• PDB: 1IVS• Seen in all Domains of Life:
• Archaea – Haloferax volcanii
• Bacteria- Escherichia Coli• Eukaryote- Homo Sapiens,
Saccharomyces cerevisiae• Important for translation of
mRNA for protein synthesis
PDB: 1IVSValine-tRNA Ligase, BRENDA Enzyme Database, http://www.brenda-enzymes.org/enzyme.php?ecno=6.1.1.9
Reaction of ValRS
• Binds L-Valine & tRNA(Val) to form Valyl-tRNA
• ATP cofactor
http://blopig.com/blog/wp-content/uploads/2013/04/Translation.jpg Modified version of http://www.bio.davidson.edu/genomics/2005/Drysdale/MFYG.html
Structure:• PDB ID: 1IVS• ValRS+AMS+tRNA(Val)• Asymmetric Homodimer
o 2 Subunitso 4 Domains per sub unit
• Large crescent shaped binding pocket
• 37 Alpha helices; 380 residues
• 42 Beta strands; 129 residues
PDB: 1IVSAsymmetric Mirror Plane
Secondary Structure:
PDB: 1IVS
Structure: ValRS+Val+AMS
ValRS
L-Valine+AMS
PDB: 1IVS
Structure:ValRS+tRNA(Val)
tRNA(Val)
ValRS
Structure:ValRS+tRNA(Val)+ATP+L-
Valine
PDB: 1IVS
Primary Structure
and Amino Acid
Sequence
Alignment
Accession Number Organism A6L3G7 Bacteroides vulgatus Q9PK91 Chlamydia muridarum Q82ZW6 Enterococcus faecalis Q74JZ8 Lactobacillus johnsonii Q4A917 Mycoplasma hyopneumoniae Q3A253 Pelobacter carbinolicus A3PHJ6 Rhodobacter sphaeroides Q7X2N3 Sphingomonas elodea P0DG64 Streptococcus pyogenes serotype M3 P46216 Trichomonas vaginalis
• Comparison of 10 Organisms
• Large amounts of conservation:
• ATP binding site• L-Valine binding site• tRNA(Val) anticodon
binding sites• tRNA(Val) binding
sites• etc
http://www.brenda-enzymes.org/sequences.php?f[stype_ec]=1&f[ec]=6.1.1.9
Interesting Conserved Amino Acids
• Lysine, Methionine, Serine, Lysine, Serine• Characteristic of Catalytic site of aminoacyl-tRNA
synthetases
Hountondji, C. et al. (2002) Crucial Role of Conserved Lysine 277 in the Fidelity of tRNA Aminoaclation by Escherichia coli Valyl-tRNA Synthetase. BioChem, 41, 14856-14865.
Lys, Met, Ser, Lys, Ser
PDB: 1IVS
• Lys528-Ser532
• L-Valine + AMS (Yellow)• tRNA (Orange)
Ser530
Lys531Lys528
Met533
Ser532
PDB: 1IVS
Binding Site:ValRS+tRNA(Val)
tRNA(Val)
Binding Site:ValRS+tRNA(Val)
PDB: 1IVS
Arg570 – tRNA Backbone interaction
CCATerminalBinding
Anti-Codon Interactions
Fukai, S. et al. (2000) Structural Basis for Double-Sieve Discrimination of L-Valine from L-Isoleucine and L-Threonine by the Comple of tRNAVal and Valyl-tRNA Synthetase. Cell, 103, 793-803.
• CCA Terminal Interactions:• C974(Grey) –side chain
interactions with:• Glu261,281 (Magenta)• Leu278 (Red)
• C975(Grey) –”Contacts with:”• Glu261 (Magenta)• Phe264 (Blue)
• A976(Grey)- H-Bond with:• Tyr337 (Green)
• A976 (Grey)-Sandwiched between:
• Leu269 (Red)• Phe264 (Blue)
Binding Site:CCA Terminal
PDB: 1IVSFukai, S. et al. (2000) Structural Basis for Double-Sieve Discrimination of L-Valine from L-Isoleucine and L-Threonine by the Comple of tRNAVal and Valyl-tRNA Synthetase. Cell, 103, 793-803.
Glu281
Leu278
Leu269
Phe264 Glu261
Tyr337
Anticodon Interactions
• Asn584 o H-Bonds with hydroxyl groups on:
• C933• A934
• A934o Side Chain interactions with:
• Phe588 (Blue)• Leu650 (Red)• Cys646 (Yellow)
• C935o Hydrogen Bonding
• N3 to Lys581• Phe588PDB: 1IVS
• Anti-Codon:• CAC• Minor Groove
Interactions
Fukai, S. et al. (2000) Structural Basis for Double-Sieve Discrimination of L-Valine from L-Isoleucine and L-Threonine by the Comple of tRNAVal and Valyl-tRNA Synthetase. Cell, 103, 793-803.
Lys581
Asn584
Phe588
Cys646
Leu650
Interesting Conserved Amino Acids
• Lysine270• Involved in editing• Edits by nucleophilic attack• Second part of the “Double Sieve”
Hountondji, C. et al. (2002) Crucial Role of Conserved Lysine 277 in the Fidelity of tRNA Aminoaclation by Escherichia coli Valyl-tRNA Synthetase. BioChem, 41, 14856-14865.
PDB: 1IVS
Lysine270 tRNA Interaction
Lys270
Lysine270 tRNA Interaction
• Lys270 (Yellow)• Up and Down stream
ValRS AA (Green)
• C975 (Red)• Down stream tRNA
Nucleic Acids (Orange)
• Interaction:o Amine group of Lys270
would preform a nucleophilic attack on improper amino acid
PDB: 1IVS
Lys270
C975
Hountondji, C. et al. (2002) Crucial Role of Conserved Lysine 277 in the Fidelity of tRNA Aminoaclation by Escherichia coli Valyl-tRNA Synthetase. BioChem, 41, 14856-14865
Double Sieve
Fukai, S. et al. (2000) Structural Basis for Double-Sieve Discrimination of L-Valine from L-Isoleucine and L-Threonine by the Comple of tRNAVal and Valyl-tRNA Synthetase. Cell, 103, 793-803.
Valine
Threonine
Isoleucine
ValRS Proofreading and Editing Pathways
Gruic-Sovulj, I. et al. (2007) Hydrolysis of non-cognate aminoacyl-adenylates by a class II aminoacyl-tRNA synthetase lacking an editing domain. FEBS Letters, 581, 26, 5110-5114
Why should we care?• Without ValRS:
• Valine would not be added to any protein sequence• This would change proteins sequence leading to:
• Non-native conformations• Malfunctioning protein• Nonfunctional protein
• Malfunctioning ValRS:• This would change proteins sequence leading to:
• Non-native conformations• Malfunctioning protein• Nonfunctional protein
• All of which could result in the death of the affected organism
Conclusion• Valyl-tRNA Synthetase is an important enzyme
o Seen in all domains of lifeo Large sections of conserved amino acids
• Structureo Asymmetric homodimero Large crescent shapeo Mainly alpha helices
• Bindingo L-Valine and ATP via KMSKSo tRNA
• Backbone• Minor groove• CCA terminal• Anti-codon
• Editingo Multiple pathways of editing pre and post transfer of tRNAo Lysine270
• Why its Important?o Accurate translation of mRNA for correct Protein sequenceo Proper conformations and functionality
Questions?
http://www.rcsb.org/pdb/education_discussion/molecule_of_the_month/images/aaRS.gif
Work Cited1 http://
www.rcsb.org/pdb/education_discussion/molecule_of_the_month/images/aaRS.gif2 PDB 1IVS3 http://blopig.com/blog/wp-content/uploads/2013/04/Translation.jpg4 http://www.bio.davidson.edu/genomics/2005/Drysdale/MFYG.html 5http://www.brenda-enzymes.org/sequences.php?f[stype_ec]=1&f[ec]=6.1.1.96 Hountondji, C.; Lazennec, C.; Beauvallet, C.; Dessen, P.; Pernollet, J.; Plateau, P.;
Blanquet, S. (2002) Crucial Role of Conserved Lysine 277 in the Fidelity of tRNA Aminoaclation by Escherichia coli Valyl-tRNA Synthetase. BioChem, 41, 14856-14865.
7 Fukai, S.; Nureki, O.; Sekine, S.; Shimada, A.; Tao, J.; Vassylyev, D.; Yokoyama, S. (2000) Structural Basis for Double-Sieve Discrimination of L-Valine from L-Isoleucine and L-Threonine by the Comple of tRNAVal and Valyl-tRNA Synthetase. Cell, 103, 793-803.
8 Gruic-Sovulj, I., Rokov-Plavec, J., Weygand-Durasevic, I. (2007) Hydrolysis of non-cognate aminoacyl-adenylates by a class II aminoacyl-tRNA synthetase lacking an editing domain. FEBS Letters, 581, 26, 5110-5114