GONUTS has been updated to MW1.31 Most things seem to be working but be sure to report problems.

Have any questions? Please email us at ecoliwiki@gmail.com

PMID:10716990

From GONUTS
Jump to: navigation, search
Citation

Yamaji, R, Adamik, R, Takeda, K, Togawa, A, Pacheco-Rodriguez, G, Ferrans, VJ, Moss, J and Vaughan, M (2000) Identification and localization of two brefeldin A-inhibited guanine nucleotide-exchange proteins for ADP-ribosylation factors in a macromolecular complex. Proc. Natl. Acad. Sci. U.S.A. 97:2567-72

Abstract

Two brefeldin A (BFA)-inhibited guanine nucleotide-exchange proteins for ADP-ribosylation factors, 200-kDa BIG1 and 190-kDa BIG2, were copurified from bovine brain cytosol associated with >670-kDa macromolecular complexes. When observed by immunofluorescence in HeLa S3 and HepG2 cells, endogenous BIG1 and coexpressed BIG2 were distributed in a punctate pattern throughout the cytosol, and also concentrated in the perinuclear region, where endogenous BIG1 and BIG2 each partially colocalized with Golgi-specific 58K protein and gamma-adaptin. On Western blot analysis, both BIG1 and BIG2 were clearly more abundant in the cytosol than in the microsomal fractions. After density gradient centrifugation of a microsomal fraction, BIG1 and BIG2 were recovered in the same fraction as beta-COP, a marker for Golgi membranes. When cytosol from HeLa S3 cells was subjected to gel filtration and fractions were analyzed by Western blotting, the largest percentages of both BIG1 and BIG2 were detected in fractions containing proteins with a molecular mass of >670 kDa. Western blotting using anti-peptide antibodies specific for BIG1 or BIG2 demonstrated that approximately 70% of BIG2 was immunoprecipitated along with 100% of BIG1 by the anti-BIG1 IgG, and approximately 75% of BIG1 was coprecipitated with 100% of BIG2 by the anti-BIG2 IgG. All observations were consistent with the conclusion that significant fractions of BIG1 and BIG2 exist as components of the same macromolecular complexes in bovine brain cytosol and are similarly localized in cultured cells.

Links

PubMed PMC15969

Keywords

ADP-Ribosylation Factors/chemistry; Blotting, Western; Brefeldin A/metabolism; Cytosol/metabolism; Dose-Response Relationship, Drug; GTP-Binding Proteins/chemistry; GTP-Binding Proteins/isolation & purification; Golgi Apparatus/metabolism; Guanine Nucleotide Exchange Factors/antagonists & inhibitors; Guanine Nucleotide Exchange Factors/metabolism; HeLa Cells; Humans; Microscopy, Fluorescence; Microsomes/metabolism; Peptides; Precipitin Tests; Protein Synthesis Inhibitors/metabolism; Time Factors; Transfection; Tumor Cells, Cultured

Significance

Annotations

Gene product Qualifier GO Term Evidence Code with/from Aspect Extension Notes Status

HUMAN:BIG2

located_in

GO:0000139: Golgi membrane

ECO:0000314: direct assay evidence used in manual assertion

C

Seeded From UniProt

complete

HUMAN:BIG2

enables

GO:0005515: protein binding

ECO:0000353: physical interaction evidence used in manual assertion

UniProtKB:Q9Y6D6

F

Seeded From UniProt

complete

HUMAN:BIG1

located_in

GO:0000139: Golgi membrane

ECO:0000314: direct assay evidence used in manual assertion

C

Seeded From UniProt

complete

HUMAN:BIG1

located_in

GO:0005829: cytosol

ECO:0000314: direct assay evidence used in manual assertion

C

Seeded From UniProt

complete

HUMAN:BIG1

enables

GO:0005515: protein binding

ECO:0000353: physical interaction evidence used in manual assertion

UniProtKB:Q9Y6D5

F

Seeded From UniProt

complete

HUMAN:BIG1

part_of

GO:0005829: cytosol

ECO:0000314: direct assay evidence used in manual assertion

C

Seeded From UniProt

complete

HUMAN:BIG1

part_of

GO:0000139: Golgi membrane

ECO:0000314: direct assay evidence used in manual assertion

C

Seeded From UniProt

complete

Notes

See also

References

See Help:References for how to manage references in GONUTS.