Structural analysis of Salmonella enterica effector protein SopD.
Salmonella outer protein D (SopD) is a type III secreted virulence effector protein from Salmonella enterica. Full-length SopD and SopD lacking 16 amino acids at the N-terminus (SopDDeltaN) have been expressed as fusions with GST in Escherichia coli, purified with a typical yield of 20-30 mg per litre of cell culture and crystallized. Biophysical characterization has been carried out mainly on SopDDeltaN. Analytical size exclusion chromatography shows that SopDDeltaN is monomeric and probably globular in aqueous solution. The secondary structure composition, calculated from the CD spectrum, is mixed (38% alpha-helix and 26% beta-strand). Sequence analysis indicates that SopD contains a coiled coil motif, as found in numerous other type III secretion system-associated proteins. This suggests that SopD has the potential for one or more heterotypic protein-protein interactions. Limited trypsin digestion of SopDDeltaN, monitored by both one-dimensional proton NMR spectroscopy and SDS-PAGE, shows that the protein has a large, protease-resistant core domain of 286 amino acid residues. This single-domain architecture suggests that SopD lacks a cognate chaperone. In crystallization trials, SopDDeltaN produced better crystals than either full-length SopD or trypsin-digested SopDDeltaN. Diffraction to 3.0 A resolution has so far been obtained from crystals of SopDDeltaN.
Wood
Michael W
MW
Division of Environmental Microbiology, Institute for Animal Health, Compton Laboratory, Berkshire RG20 7NN, UK.
Williams
Christopher
C
Upadhyay
Abhishek
A
Gill
Andrew C
AC
Philippe
Didier L
DL
Galyov
Edouard E
EE
van den Elsen
Jean M H
JM
Bagby
Stefan
S
D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D002942:Circular Dichroism; D003460:Crystallization; D019779:Salmonella enterica / Q000737:chemistry* / Q000235:genetics; D012645:Selenomethionine / Q000378:metabolism; D020539:Sequence Analysis, Protein; D017384:Sequence Deletion
D001426:Bacterial Proteins; C116105:SopD protein, Salmonella; D012645:Selenomethionine