BIG1 and BIG2, brefeldin A-inhibited guanine nucleotide-exchange factors for ADP-ribosylation factors

• Published in: Methods in enzymology Vol. 404; p. 174
• Main Authors: Jones, Heather D, Moss, Joel, Vaughan, Martha
• Format: Journal Article
• Language: English
• Published: United States 2005
• Subjects: ADP-Ribosylation Factors - metabolism; Animals; Cattle; Guanine Nucleotide Exchange Factors - metabolism; Humans; Protein Interaction Mapping
• DOI: 10.1016/s0076-6879(05)04017-6
• ISSN: 0076-6879

BIG1 and BIG2 are large (approximately 200 kDa) guanine nucleotide-exchange proteins for ADP-ribosylation factors, or ARFs, that were isolated based on sensitivity of their guanine nucleotide-exchange activity to inhibition by brefeldin A. The intracellular distributions of BIG1 and BIG2 differ from those of other ARF guanine nucleotide-exchange proteins. In addition to its presence in Golgi membranes, BIG2 is seen in peripheral vesicular structures that most likely represent recycling endosomes, and BIG1 is found in nuclei of serum-starved HepG2 cells. Several binding partners for BIG1 and BIG2 that were identified via yeast two-hybrid screens include FKBP13 and myosin IXb for BIG1 and, for BIG2, the regulatory RIalpha subunit of protein kinase A, Exo70, and the GABA receptor beta subunit. Autosomal recessive periventricular heterotopia with microcephaly, a disorder of human embryonic development due to defective vesicular trafficking, has been attributed to mutations in BIG2. Methods for purification of BIG1 and BIG2 from HepG2 cells are presented here, along with a summary of information regarding their structure and function.