Protein structure and dynamics determined by protein modeling combined with spectroscopic techniques.
Beside of the protein crystals, another attractive option in protein structure analysis has recently appeared: computer modeling of the protein structure based on homology and similarity with proteins of already known structures. We used the combination of computer modeling with spectroscopic techniques, such as steady-state or time-resolved fluorescence spectroscopy or Raman spectroscopy, and with molecular biology techniques. This method could achieve reliable results comparable with resolution obtained from crystal structures. Molecular modeling of the ATP site within the H4-H5-loop revealed eight amino acids residues, namely besides the previously reported amino acids Asp443, Lys480, Lys501, Gly502 and Arg544, also Glu446, Phe475 and Gln482, which form the complete ATP recognition site. Moreover, we proved that a hydrogen bond between Arg423 and Glu472 supported the connection of two opposite halves of the ATP-binding pocket. Similarly, the conserved residue Pro489 is important for the proper interaction of the third and fourth-strands, which both contain residues that take part in the ATP-binding (Ref. 34).
Lansky
Z
Z
Department of Protein Structure, Institute of Physiology, Academy of Sciences of the Czech Republic, Prague.
Kubala
M
M
Varga
F
F
Kolacna
L
L
Amler
E
E
D000255:Adenosine Triphosphate / Q000378:metabolism; D001665:Binding Sites; D003198:Computer Simulation; D011487:Protein Conformation; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D013059:Spectrum Analysis, Raman
D000255:Adenosine Triphosphate; D000254:Sodium-Potassium-Exchanging ATPase