Studies of the structure-function relationships of Neurospora pyruvate kinase: denaturation by urea and the effect of ligands on the refolding and renaturation process.
1. The renaturation of Neurospora pyruvate kinase following denaturation with urea was investigated. 2. The substrates, phosphoenol pyruvate and adenosine diphosphate, were observed to stabilize the enzyme against urea-induced structural disorganization. 3. It was demonstrated that for refolding/reactivation of the denatured protein the allosteric activator, fructose-1,6-diphosphate and a sulphydryl protectant are required. 4. The enzyme recovered following renaturation showed complete immunological identity with the native enzyme in Ouchterlony double diffusion tests.
D000244:Adenosine Diphosphate / Q000378:metabolism; D004789:Enzyme Activation; D008024:Ligands; D009491:Neurospora / Q000201:enzymology*; D009492:Neurospora crassa / Q000201:enzymology*; D010728:Phosphoenolpyruvate / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation / Q000187:drug effects; D011489:Protein Denaturation / Q000187:drug effects; D011770:Pyruvate Kinase / Q000378:metabolism*; D013329:Structure-Activity Relationship; D014508:Urea
D008024:Ligands; D000244:Adenosine Diphosphate; D010728:Phosphoenolpyruvate; D014508:Urea; D011770:Pyruvate Kinase