The following text describes a protein:
Function:
Receptor-binding subunit of subtilase cytotoxin SubAB5. Required for receptor-binding and thus correct trafficking in the host cell. Has specificity for host glycans terminating in the sialic acid N-glycolyl-alpha-neuraminic acid (Neu5Gc); each subunit in the SubB pentamer binds one Neu5Gc. The protease subunit (SubA) cleaves host BiP/HSPA5, inducing the host endoplasmic reticulum stress response and eventual cell death. Culture supernatant of E.coli expressing both subA and subB are toxic for Vero cells (African green monkey kidney cell line), Chinese hamster ovary cells and Hct-8 cells (human colonic epithelial cell line); the subunits are not toxic individually. Purified SubAB5 is highly toxic, <0.1 pg is able to kill at least 50% of 30'000 Vero cells in a microtiter plate assay after 3 days; no cytotoxicity is seen at 24 hours. Preabsorption with cells expressing a ganglioside GM2 mimic reduced cytotoxicity of SubAB5 by 93% in the Vero cytotoxicity assay. Intraperitoneal injection of 200 ng of purified SubAB5 kills mice; the higher the dose the faster the mice die. Animals injected with purified SubAB5 have microvascular thrombi in the brain and other organs, including the renal tubules and glomeruli. Mice fed E.coli cells expressing cloned SubAB5 experience drastic weight loss and appear ill and lethargic. SubB alone at 2.5 ug/ml causes vacuolation of Vero cells, which requires the V-type ATPase proton pump; treated cells die. Protein synthesis in Vero cells is transiently inhibited by SubAB5; both subunits are required for this effect. Inhibition of protein synthesis is prevented by brefeldin A; cells are arrested in the G1 phase. SubAB5 at 100 ng/ml induced caspase-dependent apoptosis in Vero cells through mitochondrial membrane damage.
Miscellaneous:
MISCELLANEOUS: The E.coli strain this operon was isolated from causes hemolytic uremic syndrome (HUS) and also encodes a Shigella-type toxin Stx; MISCELLANEOUS: Humans cannot convert CMP-Neu5Ac to CMP-Neu5Gc due to a mutation in the CMAHP gene, and so should be less susceptible to this toxin than other animals. However Neu5Gc can be acquired from red meat and milk products, resulting in Neu5Gc incorporation into human tissue.
Subcellular Localization:
Secreted. Host cytoplasm, host cytosol. Host endoplasmic reticulum lumen. Note=Colocalizes with host BiP/HSPA5 in the endoplasmic reticulum of Vero cells, its activity on BiP/HSPA5 is blocked by pretreatment with brefeldin A, which disrupts the Golgi apparatus and inhibits retrograde transport from the cell surface to the Golgi. Using different inhibitors it has been shown to be actively internalized by membrane-bound vesicles and undergoes clathrin-dependent retrograde transport, via early endosomes and the Golgi network, to the endoplasmic reticulum. Trafficking is similar in Vero cells, human HeLa cells and murine N2A cells.
Domain:
A patch of hydrophobic and neutral of amino acids (residues 89-94) forms a ring around the central pore of the pentamer, which interacts with the A2 helix of the SubA subunit; some mutations in this patch prevent SubAB5 complex formation but still allow some targeting to the host endoplasmic reticulum lumen. One residue in this patch, Thr-92, is the only residue that contacts SubA from all 5 of the SubB subunits.
Sequence:
MTIKRFFVCAGIMGCLSLNPAMAEWTGDARDGMFSGVVITQFHTGQIDNKPYFCIEGKQSAGSSISACSMKNSSVWGASFSTLYNQALYFYTTGQPVRIYYKPGVWTYPPFVKALTSNALVGLSTCTTSTECFGPDRKKNS