Site-specific unglycosylation to improve crystallization of the metabotropic glutamate receptor 3 extracellular domain.

ABSTRACT: Metabotropic glutamate receptors (mGluRs) are involved in the regulation of many physiological and pathological processes in the central nervous system. The extracellular domain (ECD) of mGluR subtype 3 (mGluR3) was produced using the baculovirus expression system and purified from the culture medium. However, the recombinant protein showed heterogeneity in molecular weight on SDS-PAGE analysis. It was found that the unglycosylation of Asn414 significantly reduced the heterogeneity. Consequently, three site-specifically unglycosylated mutant proteins of mGluR3 ECD, replacing Asn414 only or replacing Asn414 in combination with other glycosylation sites, were successfully crystallized in the presence of L-glutamate. Among them, crystals of the N414/439Q mutant diffracted X-rays to 2.35 A res

SUBMITTER: Muto T 

PROVIDER: S-EPMC2650456 | biostudies-literature | 2009 Mar

REPOSITORIES: biostudies-literature