Details

Autor(en) / Beteiligte

• Pardo, Miguel A.
• Llama, María J.
• Serra, Juan L.

Titel

Thermal stabilization by its ligands of NADP+-isocitrate dehydrogenase from the thermophilic cyanobacterium Phormidium laminosum

Ist Teil von

• Progress in Biotechnology, 1998, Vol.15, p.229-234

Erscheinungsjahr

1998

Link zum Volltext

Beschreibungen/Notizen

• The thermostability of the NADP+-dependent isocitrate dehydrogenase [Isocitrate: NADP+ oxidoreductase (decarboxylating), EC 1.1.1.42] purified to homogeneity from the thermophilic non-N2-fixing cyanobacterium Phormidium laminosum was studied. The catalytic activity of the enzyme increased up to 55°C and the calculated activation energy (Ea) for catalysis was 9.5 ± 0.1 kcal mol1. In the absence of ligands the enzyme was fully stable until 50°C, but more than 75% of its activity was lost when it was incubated for 15 min at 60°C, and it was completely inactive at 65°C. The thermal inactivation obeyed a first-order reaction and the calculated Ea for enzyme denaturation was 91.5 ± 0.1 kcal mol‒1. The presence of either D,L-isocitrate or NADP+ did not stabilize the enzyme, but the combined presence of D,L-isocitrate and Mn2+ increased significantly the enzyme thermostability and, at least, 50% of the activity remained after 15 min at 73°C. The calculated Ea for thermal inactivation of the enzyme-D,L-isocitrate-Mn2+ ternary complex was 109 ± 0.1 kcal mol1. These results together with the fact that the Km values of the enzyme for D,L-isocitrate and NADP+ depended on the nature of the divalent metal cation used as a cofactor suggest that the D,L-Isocitrate-metal2+ complex is the true substrate of the enzyme.