Properties of microsomal acyl-CoA: lysophosphatidylcholine acyltransferase from liver of thermally-acclimated rainbow trout,Salmo gairdneri

Abstract

Acyl-CoA: lysophosphatidylcholine acyltransferase (LPCAT) (EC 2.3.1.23) activity was assayed in liver microsomes from rainbow trout,Salmo gairdneri, acclimated to 5°C and 20°C to assess its contribution to the temperature-induced restructuring of phospholipid acyl chain composition. The synthesis of phosphatidylcholine (PC) (from lyso-PC) was threefold the synthesis of phosphatidylethanolamine (PE) (from lyso-PE) under similar assay conditions. LPCAT activity (i) displayed an absolute requirement for lysophosphatidylcholine (LPC) and was enhanced by the presence of ATP, MgCl2 and CoA (which reduced the impact of endogenous acyl-CoA hydrolase activity by regenerating the acyl-CoA substrate) in the assay medium; (ii) remained linear with time up to 30 min; and (iii) increased linearly with microsomal protein concentration up to 0.2 mg/ml for the 20°C assay and 0.4 mg/ml for the 5°C assay. There was no difference in Km or Vmax values due to the acclimation history of the fish, but there were obvious differences due to assay temperature. The apparent Km values for LPC were 58.54±7.24 μM and 12.26±2.14 μM when assayed at 5°C and 20°C respectively; values for oleoyl-CoA were 9.11±0.78 μM and 1.23±0.25 μM under the same assay conditions. Activity was 1.99±0.31 nmol min−1 mg protein−1 when assayed at 5°C, and 3.8±0.45 nmol min−1 mg protein−1 when assayed at 20°C. These findings indicate that adjustments in the activity of LPCAT play no significant role in the temperature-induced restructuring of PC molecular species composition. However, the marked temperature dependence of the Km values for LPC and oleoyl CoA suggest that patterns of fatty acid incorporation (i.e. substrate preference) may vary with assay temperature, and in this way LPCAT could contribute to the restructuring response.