Structural and nucleotide-binding properties of YajQ and YnaF, two Escherichia coli proteins of unknown function
Résumé
Structural genomics is a new approach in functional assignment of proteins identified via whole-genome sequencing programs. Its rationale is that nonhomologous proteins performing similar or related biological functions might have similar tertiary structure. We used dye pseudoaffinity chromatography, two-dimensional gel electrophoresis, and mass spectrometry to identify two novel Escherichia coli nucleotide-binding proteins, YnaF and YajQ. YnaF exhibited significant sequence identity with MJ0577, an ATP-binding protein from a hyperthermophile (Methanococcus jannaschii), and with UspA, a protein from Haemophilus influenzae that belongs to the Universal Stress Protein family. YnaF conserves the ATP-binding site and the dimeric structure observed in the crystal of MJ0577. The protein YajQ, present in many bacterial genomes, is missing in eukaryotes. In the absence of significant similarities of YajQ to any solved structure, we determined its structural and ligand-binding properties by NMR and isothermal titration calorimetry. We demonstrate that YajQ is composed of two domains, each centered on a beta-sheet, that are connected by two helical segments. NMR studies, corroborated with local sequence conservation among YajQ homologs in various bacteria, indicate that one of the beta-sheets is mostly involved in biological activity.
Mots clés
Adenosine Triphosphate
Amino Acid Sequence
Calorimetry
Circular Dichroism
Dimerization
Escherichia coli Proteins
Ligands
Models
Molecular
Molecular Sequence Data
Molecular Structure
Nuclear Magnetic Resonance
Biomolecular
Nucleotides
Protein Binding
Protein Conformation
Protein Structure
Secondary
Tertiary
Proteome
Recombinant Proteins
RNA-Binding Proteins
Sequence Alignment