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Eukaryotic expression, purification, crystallization and preliminary X-ray analysis of murine Manic Fringe.

ABSTRACT: Fringe proteins are Golgi-resident beta1,3-N-acetylglucosaminyltransferases that regulate development in metazoa through glycosylation of the Notch receptor and its ligands. The catalytic domain of murine Manic Fringe was expressed in the baculovirus/insect-cell system as a secreted protein. Mass-spectrometric analysis of the purified protein indicated the presence of two N-linked glycans. Abolishing the glycosylation sites by site-directed mutagenesis was necessary in order to obtain orthorhombic crystals that diffracted to 1.8 angstroms resolution. For phasing, a highly redundant data set was collected using a crystal soaked with halide salts.

SUBMITTER: Jinek M

PROVIDER: S-EPMC2242911 | biostudies-literature | 2006 Aug

REPOSITORIES: biostudies-literature

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Eukaryotic expression, purification, crystallization and preliminary X-ray analysis of murine Manic Fringe.

Jinek Martin M Conti Elena E

Acta crystallographica. Section F, Structural biology and crystallization communications 20060724 Pt 8

Fringe proteins are Golgi-resident beta1,3-N-acetylglucosaminyltransferases that regulate development in metazoa through glycosylation of the Notch receptor and its ligands. The catalytic domain of murine Manic Fringe was expressed in the baculovirus/insect-cell system as a secreted protein. Mass-spectrometric analysis of the purified protein indicated the presence of two N-linked glycans. Abolishing the glycosylation sites by site-directed mutagenesis was necessary in order to obtain orthorhomb ...[more]

PMID: 16880554

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