PRODUCTION OF BIOLOGICALLY-ACTIVE RECOMBINANT AVIDIN IN BACULOVIRUS-INFECTED INSECT CELLS

An efficient lepidopteran insect cell system was established for the e xpression of a recombinant form of chicken egg-white avidin. The gene product was obtained in both secreted and intracellular forms, and bio logically active recombinant avidin was isolated using affinity chroma tography on an iminobiotin-agarose column. Similar to the known quater nary structure of the native egg-white protein, the purified recombina nt protein was glycosylated and assembled mainly into tetramers. Like native avidin, the recombinant tetramer also exhibited a high level of thermostability, and was further stabilized upon binding biotin. The biotin-binding and structural properties of the recombinant avidin are thus similar to those of the natural egg-white protein, and the insec t system is appropriate both for future site-directed mutagenesis stud ies and for the production of avidin fusion proteins. (C) 1997 Academi c Press.