Opposing effects of two osmolytes--trehalose and glycerol--on thermal inactivation of rabbit muscle 6-phosphofructo-1-kinase.
Trehalose and glycerol are known as good stabilizers of function and structure of several macromolecules against stress conditions. We previously reported that they have comparable effectiveness on protecting two yeast cytosolic enzymes against thermal inactivation. However, enzyme protection has always been associated to a decrease in catalytic activity at the stabilizing conditions i.e., the presence of the protective molecule. In the present study we tested trehalose and glycerol on thermal protection of the mammalian cytosolic enzyme phosphofructokinase. Here we found that trehalose was able to protect phosphofructokinase against thermal inactivation as well as to promote an activation of its catalytic activity. The enzyme incubated in the presence of 1 M trehalose did not present any significant inactivation within 2 h of incubation at 50 degrees C, contrasting to control experiments where the enzyme was fully inactivated during the same period exhibiting a t0.5 for thermal inactivation of 56+/-5 min. On the other hand, enzyme incubated in the presence of 37.5% (v/v) glycerol was not protected against incubation at 50 degrees C. Indeed, when phosphofructokinase was incubated for 45 min at 50 degrees C in the presence of lower concentrations of glycerol (7.5-25%, v/v), the remaining activity was 2-4 times lower than control. These data show that the compatibility of effects previously shown for trehalose and glycerol with some yeast cytosolic enzymes can not be extended to all globular enzyme system. In the case of phosphofructokinase, we believe that its property of shifting between several different complex oligomers configurations can be influenced by the physicochemical properties of the stabilizing molecules.
Faber-Barata
Joana
J
Laboratório de Enzimologia e Controle do Metabolismo, Departamento de Fármacos, Faculdade de Farmácia, Universidade Federal do Rio de Janeiro, Rio de Janeiro, RJ, Brasil.
Sola-Penna
Mauro
M
D000818:Animals; D002384:Catalysis / Q000187:drug effects; D004789:Enzyme Activation; D004795:Enzyme Stability; D005990:Glycerol / Q000737:chemistry / Q000494:pharmacology*; D006358:Hot Temperature; D025203:Phosphofructokinase-1, Muscle Type / Q000737:chemistry* / Q000187:drug effects*; D011817:Rabbits; D014199:Trehalose / Q000737:chemistry / Q000494:pharmacology*
D014199:Trehalose; D025203:Phosphofructokinase-1, Muscle Type; D005990:Glycerol