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Research Themes Cell biology

Terminal activation

SBKB [doi:10.1038/sbkb.2011.51]
Featured Article - December 2011
Short description: The recent crystal structure of the human BIG2 Sec7 domain reveals a novel ARF-interacting motif that regulates ARF activation.

Model of human BIG2 Sec7 domain (PDB 3L8N, green) in a complex with ARF1 (cyan). The loop>J of BIG2 Sec7 domain is positioned close to the switch I (Sw. I) region of ARF1. Image courtesy of Elizabeth Sztul.

G proteins are key signaling molecules that serve as molecular switches by cycling between an inactive GDP-bound state and an active GTP-bound state. G proteins are activated by guanine nucleotide exchange factors (GEFs), which catalyze the exchange of GDP for GTP, and inactivated by GTPase activating proteins (GAPs), which catalyze the hydrolysis of GTP to GDP. Understanding the interplay between G proteins, GEFs, and GAPs is essential to discovering the intricate signaling pathways in the cell.

As part of the PSI:Biology's Community Outreach activities, Sztul and colleagues with collaborators from the PSI NESG have examined the activation of ADP-ribosylation factors (ARFs), a family of G proteins involved in membrane trafficking and intracellular signaling, by GEFs. ARF GEFs are characterized by a conserved Sec7 domain, which is responsible for ARF binding and for catalyzing the exchange of GDP for GTP. The authors determined the crystal structure of the Sec7 domain of the ARF GEF human BIG2 which, along with other Sec7 domain family members, is a target of the NESG Human Cancer Protein Interaction program. The structure, which is similar to structures of other Sec7 domains, reveals an elongated rod-shaped protein consisting of ten α-helices with a hydrophobic groove forming the ARF-binding interface. Upon modeling the ARF-Sec7 domain complex, the authors predicted a novel ARF-interacting motif in the C-terminus of the Sec7 domain located in the loop after helix J (loop>J).

The authors investigated the functional role of loop>J by mutating residues in loop>J from Sec7 domains of several ARF GEFs (BIG2, GBF1, and ARNO). Mutations in loop>J in BIG2 and GBF1 produced phenotypes similar to catalytically dead mutants in vivo, suggesting that loop>J is involved in catalyzing guanine nucleotide exchange. Pull-down and FRAP assays with BIG2, GBF1, and ARNO revealed that mutation of loop>J decreases ARF binding, resulting in a decrease in GDP to GTP exchange.

This is the first study to identify the role of loop>J in ARF GEFs in substrate recognition and catalysis. Understanding the interactions that lead to ARF activation could provide insight into the selectivity of ARF GEFs and aid in designing GEF-specific inhibitors.

Jennifer Cable

References

  1. J. Lowery et al. Novel C-terminal Motif within Sec7 Domain of Guanine Nucleotide Exchange Factors Regulates ADP-ribosylation Factor (ARF) Binding and Activation.
    J. Biol. Chem. 286, 36898-36906 (2011). doi:10.1074/jbc.M111.230631

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