The structure of RalF, an ADP-ribosylation factor guanine nucleotide exchange factor from Legionella pneumophila, reveals the presence of a cap over the active site.
The Legionella pneumophila protein RalF is secreted into host cytosol via the Dot/Icm type IV transporter where it acts to recruit ADP-ribosylation factor (Arf) to pathogen-containing phagosomes in the establishment of a replicative organelle. The presence in RalF of the Sec7 domain, present in all Arf guanine nucleotide exchange factors, has suggested that recruitment of Arf is an early step in pathogenesis. We have determined the crystal structure of RalF and of the isolated Sec7 domain and found that RalF is made up of two domains. The Sec7 domain is homologous to mammalian Sec7 domains. The C-terminal domain forms a cap over the active site in the Sec7 domain and contains a conserved folding motif, previously observed in adaptor subunits of vesicle coat complexes. The importance of the capping domain and of the glutamate in the "glutamic finger," conserved in all Sec7 domains, to RalF functions was examined using three different assays. These data highlight the functional importance of domains other than Sec7 in Arf guanine nucleotide exchange factors to biological activities and suggest novel mechanisms of regulation of those activities.
Amor
J Carlos
JC
Department of Biochemistry, Emory University School of Medicine, Atlanta, Georgia 30322, USA.
Swails
Jennifer
J
Zhu
Xinjun
X
Roy
Craig R
CR
Nagai
Hiroki
H
Ingmundson
Alyssa
A
Cheng
Xiaodong
X
Kahn
Richard A
RA
D020727:ADP-Ribosylation Factors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D020662:Guanine Nucleotide Exchange Factors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D016952:Legionella pneumophila / Q000166:cytology / Q000201:enzymology* / Q000235:genetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000378:metabolism; D013329:Structure-Activity Relationship; D014617:Vacuoles / Q000378:metabolism
D001426:Bacterial Proteins; D020662:Guanine Nucleotide Exchange Factors; D011506:Proteins; C066755:Sec7 guanine nucleotide exchange factors; D006160:Guanosine Triphosphate; D020727:ADP-Ribosylation Factors