The sequential nature of the negative cooperativity in rabbit muscle glyceraldehyde-3-phosphate dehydrogenase.
The binding of nicotinamide--adenine dinucleotide (NAD+), nicotinamide--1,N6-ethenoadenine dinucleotide (epsilon NAD+), acetylpyridine--adenine dinucleotide (AcPyAD+), ATP, and adenosine diphosphoribose (ADP-ribose) to rabbit muscle glyceraldehyde-3-phosphate dehydrogenase (the enzyme) was examined. NAD+ and epsilon NAD+ were found to bind to the apoenzyme in a negatively cooperative manner, whereas AcPyAD+, ATP, and ADP-ribose bind non-cooperatively to the NAD+ sites. The strong negative cooperativity in coenzyme binding was found to be abolished in the presence of AcPyAD+ and strongly weakened by ATP, ADP, and AMP, but was not affected by the addition of ADP-ribose. These findings demonstrate that the mechanism of the negative cooperativity in coenzyme binding to the enzyme involves ligand-induced conformational changes between neighboring sites. These findings cannot be accounted for by the pre-existent asymmetry model. The results support our previous hypothesis that the structure of the pyridine moiety of the coenzyme analogues plays a role in orienting the adenine moiety in the adenine subsite, and thus affects the cooperativity observed in the binding of the coenzyme analogue.
D000246:Adenosine Diphosphate Ribose / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001667:Binding, Competitive; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000378:metabolism*; D007700:Kinetics; D046911:Macromolecular Substances; D009132:Muscles / Q000201:enzymology*; D009243:NAD / Q000031:analogs & derivatives / Q000378:metabolism; D011817:Rabbits; D013050:Spectrometry, Fluorescence
D046911:Macromolecular Substances; D009243:NAD; D000246:Adenosine Diphosphate Ribose; C011038:nicotinamide 1,N(6)-ethenoadenine dinucleotide; C005941:3-acetylpyridine adenine dinucleotide; D000255:Adenosine Triphosphate; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases