Effect of glucose on the conformation of ADPMg(II) bound at the active site of yeast hexokinase PI.
The conformation of ADPMg(II) bound at the active site of yeast hexokinase PI has been determined using transferred nuclear Overhauser effect spectroscopy (TRNOESY). We have measured the time dependent NOE buildup of all the proton pairs of ADP in enzyme. ADPMg(II) and enzyme. glucose.ADPMg(II) complexes at 500 MHz and 10 degrees C. The data have been analyzed using complete relaxation matrix approach to obtain various inter-proton distances. These distances were used as restraints in the molecular dynamics and energy minimization to obtain the conformation of the bound nucleotide. The results from these calculations suggest that in both the complexes, the nucleotide binds in an anti conformation with a glycosidic torsion angle chi = 55 +/- 5 degrees and 52 +/- 5 degrees in PI.ADPMg(II) and PI.glucose.ADPMg(II) complexes, respectively. However, the phase angle of pseudorotation (P) which defines the sugar pucker in the two complexes was found to be 99 degrees, corresponding to 0T1 for PI.ADPMg(II) complex and 69 degrees corresponding to 4T0 for PI.glucose.ADPMg(II) complex. The cleft closure conformational change in the enzyme induced by glucose seems to affect the conformation of the ribosyl moiety of the bound nucleotide.
Maity
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Tata Institute of Fundamental Research, Colaba, Mumbai, India.
Jarori
G K
GK
D000255:Adenosine Triphosphate / Q000737:chemistry* / Q000378:metabolism*; D005947:Glucose / Q000494:pharmacology*; D006593:Hexokinase / Q000737:chemistry* / Q000378:metabolism*; D007700:Kinetics; D008958:Models, Molecular; D008968:Molecular Conformation; D019906:Nuclear Magnetic Resonance, Biomolecular; D011487:Protein Conformation / Q000187:drug effects*; D012441:Saccharomyces cerevisiae / Q000201:enzymology*
D000255:Adenosine Triphosphate; D006593:Hexokinase; D005947:Glucose