ENGINEERING OF FUNCTIONAL CHIMERIC PROTEIN-G - VARGULA LUCIFERASE

Luciferase of Vargula hilgendorfii is infinitely stable at room temper ature in dried state, and its light-emitting reaction is very simple. These unique characteristics of Vargula luciferase have prompted us to engineer chimeric protein, the other moiety chosen for conjugation be ing streptococcal protein G. A single domain of protein G which binds to IgG of a wide range of species was fused at the N-terminal region o f Vargula luciferase. Unexpectedly, we found that the chimeric protein expressed in mammalian COS-1 cells had no IgG-binding ability, probab ly due to some sort of interaction between the two moieties or some co nformational preferences of the IgG-binding domain of protein G when f used to Vargula luciferase, Here we report how we regained the IgG bin ding of protein G, by the intervention of three alpha-helices of prote in A between protein G and luciferase, To our knowledge, the new chime ric protein provides the first reported model of this kind. (C) 1997 A cademic Press.