The two-step reversible denaturation of lactate dehydrogenase at low pH.
Upon exposure to conditions of low pH, beef B4 lactate dehydrogenase rapidly loses enzymatic activity, but this process can be completely reversed yielding 100% of the original activity if the enzyme is immediately returned to neutral conditions. As the time of exposure to low pH is increased, the fraction of activity recovered declines to a values of 50--60% and remains nearly constant over a period of many hours. Correlated with this behavior is a change in the kinetics of the recovery of activity. Recovery of activity has been shown to be a second-order process for enzyme exposed to low pH for brief periods of time (Anderson, S., and Weber, G. (1966), Arch. Biochem. Biophys. 116, 207). After several minutes at low pH recovery of activity is found to become first order and to occur at a considerably slower rate. Gel filtration chromatography at low pH separates the protein into two fractions. The lower molecular weight fraction is found to be primarily monomeric, as indicated by equilibrium ultracentrifugation, and is capable of recovering enzymatic activity. The higher molecular weight fraction is generated from the lower molecular weight fraction, and is incapable of recovering activity. These results are interpreted to indicate that the enzyme exists sequentially in three denatured forms at low pH, the first two capable of being restored to the native state, and the third irreversibly denatured.
D000818:Animals; D001665:Binding Sites; D002850:Chromatography, Gel; D006863:Hydrogen-Ion Concentration; D007527:Isoenzymes; D007700:Kinetics; D007770:L-Lactate Dehydrogenase; D007834:Lasers; D009206:Myocardium / Q000201:enzymology; D011485:Protein Binding; D011489:Protein Denaturation; D013050:Spectrometry, Fluorescence; D013997:Time Factors; D014461:Ultracentrifugation
D007527:Isoenzymes; D007770:L-Lactate Dehydrogenase