Christian Radauer <rc150295 at emb1.bcc.univie.ac.at> wrote:
>Hi all,
>I am trying to express a single chain T cell receptor (i.e. my insert
>consists of alpha-chain/linker/beta-chain; together about 700 bases) in
>sf9 insect cells using the baculovirus expression system obtained from
>Pharmingen. My vector (pAcSecG2T) is designed to enable the protein to be
>secreted into the medium. I found the protein (via western blot) in the
>cell lysate (the molecular weight was correct) but not in the cell
>culture supernatant.It seems as if the protein plus leader got stuck in
>the ER due to wrong folding. Has anyone of you who works with the baculo
>expression system and tries to express heterodimers like antibodies,TCR`s,
>MHC`s,...(as single-chains or alpha and beta-chain separately) ever
>encountered a problem like this and what did you do to overcome it? I am
>eagerly awaiting some helpful hints to solve my problem.
I ain't no expert. But would these insect cells have the necessary
chaperones needed for correct protein folding?
George M. Carter
