Download Molecular Orbital Interactions in the Anticodon of Transfer RNA

Molecular Orbital Interactions in
the Anticodon of Transfer RNA
Nick H. Hopkins and Elizabeth E. Koballa
Advisors: Dr. Maria Nagan and Dr. Eric Patterson
Truman State University, Kirksville, MO
Central Dogma of Molecular Biology
Genetic information is
stored in DNA and
passed on sequentially
to RNA and then
protein synthesis.
During protein synthesis, tRNA anticodons are
matched to mRNA codons and the correct amino
acid is incorporated into the growing protein chain.
“One way street” to transfer genetic information:
DNA to mRNA to protein.
Proteins
Essential to cellular function;
the communication, transport,
and chemical reactions
Enzymatic - catalyze chemical
reactions
Structural - support
Transport - vehicle for
particle movement
(e.g., transmembrane)
Receptor - cell
recognition and particle
reception
RNA and Nucleotides
Three types - rRNA, mRNA, and tRNA
RNA is made of a nucleotide chain
Three parts of a
nucleotide phosphate, sugar
(ribose), and a
nitrogenous base
Four common
bases in RNA - A,
G, C and U
tRNA
Translates mRNA
information from codon into
specific amino acids
Carries amino acids to
ribosomes to incorporate
into polypeptide chain
73-93 nucleotides long
Five regions - D loop, TCloop, acceptor stem,
variable arm, and anticodon
Anticodon - reads mRNA
codon to dictate amino acid
tRNA from S. cerevisiae or Baker’s Yeast
Anticodon-Codon Recognition
Three anticodon
nucleotides must be in a
stair-stepped conformation
to read three mRNA codon
nucleotides in the context of
the ribosome
Anticodon= Positions 34-36
Anticodon bases of a tRNA (red, anticodon components
in silver) reading codon bases on mRNA (green). Note
the stair-stepped base stacking of the tRNA anticodon.
Anticodon Stair-Stepped Conformation
Free tRNA anticodon
exhibits stair-stepped
conformation.
Stabilizing forces in
anticodon stairstepped conformation
are unclear
Molecular orbital
interactions between
nucleotides of
consecutive bases
are examined
Modified Bases
Human tRNALys,3
Naturally occurring
modified bases found
in tRNA and required
for recognition of
mRNA
From molecular
dynamics simulations,
the base modification
at the 37th position
required for stairstepped conformation
in tRNALys,3
Goals
To generally understand the
stabilization of the tRNA anticodon
through the molecular orbital
interactions.
To quantify the specific interactions of
the stair-stepped conformation.
To determine how modified bases in
general at the 37th position add
additional stability.
Strategy
Examine molecular orbitals and the
interactions between bases in x-ray crystal
and NMR structures of anticodons
Extend analysis to systems with included
modified bases at the 37th position.
Methods
Structures of tRNA obtained from Protein
Database, a bank of structures obtained by
x-ray crystallography or NMR.
tRNA
structures edited to bases 34-36
(10 structures) or 34-37 (6 structures)
Addition of hydrogen atoms to the tRNA
molecules
Optimization of hydrogen atomic positions
with Hartree-Fock level of theory and a
3-21G basis set (~700 basis functions).
Density Functional Theory
Intended level of theory for analysis: M05-2X
Developed by the Truhlar group of U of MN
Accurate with non-bonded interactions
Available level of theory for testing: MPWB1K
Where occupied molecular orbitals interact,
the energy of the interaction is calculated.
The calculated energy can be used to
quantize the interactions and further classify
them.
Natural Bond Orbital Analysis
From molecular orbital
calculations, natural bond
orbital (NBO) analysis
calculates the contributions
of traditional atomic and
hybridized orbitals to the MO
picture.
Perturbation theory energy
analysis
Steric analysis
The highest occupied molecular orbital of
water as predicted by NBO analysis.
Results
First approach: Single point calculations
with MPWB1K/6-31+G* were carried out.
Half of the NBO with MPWB1K/6-31+G*
crashed.
Successful NBO outputs were examined to
identify favorable interactions.
Those interactions with relatively high
energy (~2 kcal/mol) were visualized with
GaussView to find patterns in the
interactions amongst tRNAs.
Future Directions
Release of the newest Gaussian
revision (03M) will allow us to perform
NBO analysis with M05-2X/6-31+G*.
This will be Elizabeth’s summer project.
Acknowledgements
Dr. Maria Nagan, Research Advisor
Dr. Eric Patterson, Research Advisor
Oscar A. McCrate (graduate student, Stanford)
Petroleum Research Fund (41701-GB7)
National Science Foundation
(RUI:CHE-0521063; MRI: CHE-0746096)