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Phenylalanyl-tRNA synthetase interacts with DNA: studies on activity using deoxyribooligonucleotides
K A Ivanov1, N A Moor, V N Ankilova
1Novosibirsk Institute of Bioorganic Chemistry, Siberian Branch of the Russian Academy of Sciences, Novosibirsk, 630090, Russia. ivkon@niboch.nsc.ru
Biochemistry. Biokhimiia
|May 16, 2000
Summary
Phenylalanyl-tRNA synthetase binds single-stranded DNA more effectively than double-stranded DNA, with binding affinity increasing for longer DNA strands. This interaction occurs near the protein
Area of Science:
- Biochemistry
- Molecular Biology
- Structural Biology
Background:
- Phenylalanyl-tRNA synthetase (PheRS) is crucial for protein synthesis, attaching phenylalanine to its cognate tRNA.
- While primarily known for aminoacylation, some synthetases exhibit moonlighting functions, including DNA binding.
- Understanding DNA-protein interactions is vital for comprehending cellular processes beyond canonical functions.
Purpose of the Study:
- To investigate the interaction between Phenylalanyl-tRNA synthetase from *Thermus thermophilus* and single-stranded (ssDNA) versus double-stranded DNA (dsDNA).
- To determine the effect of DNA length and structure on PheRS binding.
- To map the DNA binding site on the PheRS enzyme.
Main Methods:
- Gel electrophoresis and surface plasmon resonance to assess binding affinities.
- Affinity labeling techniques to identify the DNA binding site.
- Comparative analysis of *Thermus thermophilus* PheRS and *E. coli* PheRS DNA binding.
Main Results:
- PheRS from *T. thermophilus* exhibits higher affinity for short ssDNA fragments (<30 nucleotides) compared to dsDNA.
- Binding efficiency significantly increases with ssDNA length beyond 20 nucleotides.
- Affinity labeling localized the DNA binding site to the interface of the alpha- and beta-subunits, distinct from the tRNA binding site.
- Phenylalanyl-tRNA synthetase from *E. coli* was also found to bind DNA.
Conclusions:
- *Thermus thermophilus* PheRS preferentially binds to ssDNA, with affinity modulated by length.
- The DNA binding site is spatially separated from the tRNA binding site, suggesting distinct functional roles.
- The ability of PheRS from different species (*T. thermophilus* and *E. coli*) to bind DNA indicates a conserved, potentially non-canonical function.