Increased production of active human beta(2)-adrenergic/G(alpha S) fusion receptor in Sf-9 cells using nutrient limiting conditions

Using the baculovirus/insect-cell expression vector system, we succeeded in obtaining a high yield of active human beta (2)-adrenergic receptor/G(alph as) fusion protein. This was achieved following high cell density productio n under nutrient-limiting conditions using a very low multiplicity of infec tion (MOI). This approach was found to significantly reduce inactive protei n accumulation that occurred when production was done using conventional hi gh MOI procedures. The maximum specific and volumetric yields of active rec eptor using this strategy increased by factors of two- and sixfold, respect ively. Our results suggest that the increase in the ratio of active/total p rotein produced results from production under nutrient limitation. Since lo w multiplicity of infection offers many advantages for large-scale applicat ions, we suggest that this simple production method should be considered wh en optimizing expression of G-protein-coupled receptors and other complex p roteins. (C) 2001 Academic Press.