Folding intermediates of hyperthermophilic D-glyceraldehyde-3-phosphate dehydrogenase from Thermotoga maritima are trapped at low temperature.
D-Glyceraldehyde-3-phosphate dehydrogenase from the hyperthermophilic eubacterium, Thermotoga maritima, is extremely thermostable showing a thermal transition beyond 105 degrees C. At low temperature, 'cold denaturation' becomes detectable only in the presence of destabilizing agents. Reconstitution after preceding denaturation depends on temperature. At 0 degree C, no significant recovery of activity is detectable, whereas between 30 and 100 degrees C reactivation reaches up to 85%. Shifting the temperature from low values to the range of optimum reconstitution releases the trapped intermediate in a fast reaction. Evidence from ultra-centrifugal analysis and far-UV circular dichroism proves the intermediate to be partially assembled to the tetramer, with most of its native secondary structure restored in a fast reaction. Fluorescence emission exhibits at least biphasic kinetics with the rate-limiting step(s) reflecting local adjustments of aromatic residues involved in tertiary contacts in the native state of the enzyme.
Institut für Biophysik und Physikalische Biochemie, Universität Regensburg, Germany.
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D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000737:chemistry*; D006089:Gram-Negative Anaerobic Bacteria / Q000201:enzymology*; D007700:Kinetics; D011489:Protein Denaturation; D013696:Temperature; D013816:Thermodynamics; D014508:Urea / Q000737:chemistry
D014508:Urea; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases