SUBA_ECOLX
ID SUBA_ECOLX Reviewed; 347 AA.
AC Q6EZC2;
DT 28-MAR-2018, integrated into UniProtKB/Swiss-Prot.
DT 16-AUG-2004, sequence version 1.
DT 03-AUG-2022, entry version 80.
DE RecName: Full=Subtilase cytotoxin subunit A {ECO:0000303|PubMed:15226357};
DE EC=3.4.21.- {ECO:0000269|PubMed:17024087};
DE Flags: Precursor;
GN Name=subA {ECO:0000303|PubMed:15226357};
OS Escherichia coli.
OG Plasmid megaplasmid pO113.
OC Bacteria; Proteobacteria; Gammaproteobacteria; Enterobacterales;
OC Enterobacteriaceae; Escherichia.
OX NCBI_TaxID=562;
RN [1]
RP NUCLEOTIDE SEQUENCE [GENOMIC DNA].
RC STRAIN=O113:H21 / 98NK2 / STEC; PLASMID=megaplasmid pO113;
RX PubMed=11598075; DOI=10.1128/iai.69.11.6999-7009.2001;
RA Paton A.W., Srimanote P., Woodrow M.C., Paton J.C.;
RT "Characterization of Saa, a novel autoagglutinating adhesin produced by
RT locus of enterocyte effacement-negative Shiga-toxigenic Escherichia coli
RT strains that are virulent for humans.";
RL Infect. Immun. 69:6999-7009(2001).
RN [2]
RP NUCLEOTIDE SEQUENCE [GENOMIC DNA], FUNCTION, AND SUBCELLULAR LOCATION.
RC STRAIN=O29;
RX PubMed=17101670; DOI=10.1128/iai.01336-06;
RA Morinaga N., Yahiro K., Matsuura G., Watanabe M., Nomura F., Moss J.,
RA Noda M.;
RT "Two distinct cytotoxic activities of subtilase cytotoxin produced by
RT Shiga-toxigenic Escherichia coli.";
RL Infect. Immun. 75:488-496(2007).
RN [3]
RP PROTEIN SEQUENCE OF N-TERMINUS, FUNCTION, PUTATIVE ACTIVE SITE, SUBUNIT,
RP SUBCELLULAR LOCATION, OPERON, DISRUPTION PHENOTYPE, BIOTECHNOLOGY, AND
RP MUTAGENESIS OF SER-272.
RC STRAIN=O113:H21 / 98NK2 / STEC; PLASMID=megaplasmid pO113;
RX PubMed=15226357; DOI=10.1084/jem.20040392;
RA Paton A.W., Srimanote P., Talbot U.M., Wang H., Paton J.C.;
RT "A new family of potent AB(5) cytotoxins produced by Shiga toxigenic
RT Escherichia coli.";
RL J. Exp. Med. 200:35-46(2004).
RN [4]
RP ERRATUM OF PUBMED:15226357.
RX DOI=10.1084/jem.20040392111204c;
RA Paton A.W., Srimanote P., Talbot U.M., Wang H., Paton J.C.;
RT "A new family of potent AB(5) cytotoxins produced by Shiga toxigenic
RT Escherichia coli.";
RL J. Exp. Med. 200:1525-1525(2004).
RN [5]
RP SUBCELLULAR LOCATION.
RX PubMed=18042253; DOI=10.1111/j.1462-5822.2007.01085.x;
RA Chong D.C., Paton J.C., Thorpe C.M., Paton A.W.;
RT "Clathrin-dependent trafficking of subtilase cytotoxin, a novel AB5 toxin
RT that targets the endoplasmic reticulum chaperone BiP.";
RL Cell. Microbiol. 10:795-806(2008).
RN [6]
RP FUNCTION, SUBCELLULAR LOCATION, AND MUTAGENESIS OF SER-272.
RX PubMed=18005237; DOI=10.1111/j.1462-5822.2007.01094.x;
RA Morinaga N., Yahiro K., Matsuura G., Moss J., Noda M.;
RT "Subtilase cytotoxin, produced by Shiga-toxigenic Escherichia coli,
RT transiently inhibits protein synthesis of Vero cells via degradation of BiP
RT and induces cell cycle arrest at G1 by downregulation of cyclin D1.";
RL Cell. Microbiol. 10:921-929(2008).
RN [7]
RP FUNCTION, AND MUTAGENESIS OF SER-272.
RX PubMed=18433465; DOI=10.1111/j.1462-5822.2008.01164.x;
RA Wolfson J.J., May K.L., Thorpe C.M., Jandhyala D.M., Paton J.C.,
RA Paton A.W.;
RT "Subtilase cytotoxin activates PERK, IRE1 and ATF6 endoplasmic reticulum
RT stress-signalling pathways.";
RL Cell. Microbiol. 10:1775-1786(2008).
RN [8]
RP FUNCTION, AND MUTAGENESIS OF SER-272.
RX PubMed=19380466; DOI=10.1128/iai.01510-08;
RA Matsuura G., Morinaga N., Yahiro K., Komine R., Moss J., Yoshida H.,
RA Noda M.;
RT "Novel subtilase cytotoxin produced by Shiga-toxigenic Escherichia coli
RT induces apoptosis in Vero cells via mitochondrial membrane damage.";
RL Infect. Immun. 77:2919-2924(2009).
RN [9] {ECO:0007744|PDB:2IY9}
RP X-RAY CRYSTALLOGRAPHY (1.80 ANGSTROMS), DISULFIDE BONDS, FUNCTION,
RP SUBSTRATE SPECIFICITY, CATALYTIC ACTIVITY, SUBCELLULAR LOCATION, DOMAIN,
RP AND MUTAGENESIS OF SER-272.
RX PubMed=17024087; DOI=10.1038/nature05124;
RA Paton A.W., Beddoe T., Thorpe C.M., Whisstock J.C., Wilce M.C.,
RA Rossjohn J., Talbot U.M., Paton J.C.;
RT "AB5 subtilase cytotoxin inactivates the endoplasmic reticulum chaperone
RT BiP.";
RL Nature 443:548-552(2006).
RN [10] {ECO:0007744|PDB:4BWG}
RP X-RAY CRYSTALLOGRAPHY (2.60 ANGSTROMS) IN COMPLEX WITH SUBB, SUBUNIT,
RP SUBCELLULAR LOCATION, DOMAIN, AND DISULFIDE BONDS.
RX PubMed=23921389; DOI=10.1074/jbc.m113.462622;
RA Le Nours J., Paton A.W., Byres E., Troy S., Herdman B.P., Johnson M.D.,
RA Paton J.C., Rossjohn J., Beddoe T.;
RT "Structural basis of subtilase cytotoxin SubAB assembly.";
RL J. Biol. Chem. 288:27505-27516(2013).
CC -!- FUNCTION: Protease subunit of subtilase cytotoxin SubAB5
CC (PubMed:17024087). An endoprotease specific for host endoplasmic
CC reticulum (ER) chaperone BiP/HSPA5, has no activity on human HSP70 or
CC HSPA8 (PubMed:17024087). Cleaves between 'Leu-416' and 'Leu-417' of
CC BiP/HSPA5 in the hinge between BiP's ATPase and protein-binding domains
CC (PubMed:17024087). This induces host ER stress response and eventual
CC cell death (PubMed:18005237, PubMed:18433465). Culture supernatant of
CC E.coli expressing both subA and subB are toxic for Vero cells (African
CC green monkey kidney cell line), Chinese hamster ovary cells and Hct-8
CC cells (human colonic epithelial cell line); the subunits are not toxic
CC individually (PubMed:15226357). Purified SubAB5 is highly toxic, <0.1
CC pg is able to kill at least 50% of 30'000 Vero cells in a microtiter
CC plate assay after 3 days; no cytotoxicity is seen at 24 hours
CC (PubMed:15226357). Preabsorption with cells expressing a ganglioside
CC GM2 mimic reduced cytotoxicity of SubAB5 by 93% in the Vero
CC cytotoxicity assay (PubMed:15226357). Intraperitoneal injection of 200
CC ng of purified SubAB5 kills mice; the higher the dose the faster the
CC mice die. Animals injected intraperitoneally with purified SubAB5 have
CC microvascular thrombi in the brain and other organs, including the
CC renal tubules and glomeruli (PubMed:15226357). Injection induces an
CC unfolded response in mice (PubMed:17024087). Mice fed E.coli cells
CC expressing cloned SubAB5 experience drastic weight loss and appear ill
CC and lethargic (PubMed:15226357). Protein synthesis in Vero cells is
CC transiently inhibited by SubAB5; both subunits are required for this
CC effect (PubMed:17101670, PubMed:18005237, PubMed:18433465). Inhibition
CC of protein synthesis is prevented by brefeldin A; cells are arrested in
CC the G1 phase (PubMed:18005237). SubAB5 at 100 ng/ml induced caspase-
CC dependent apoptosis in Vero cells through mitochondrial membrane damage
CC (PubMed:19380466). {ECO:0000269|PubMed:15226357,
CC ECO:0000269|PubMed:17024087, ECO:0000269|PubMed:17101670,
CC ECO:0000269|PubMed:18005237, ECO:0000269|PubMed:18433465,
CC ECO:0000269|PubMed:19380466}.
CC -!- SUBUNIT: Forms a complex with SubB with the stoichiometry SubA1:SubB5
CC (called SubAB5) (PubMed:15226357, PubMed:23921389).
CC {ECO:0000269|PubMed:23921389, ECO:0000305|PubMed:15226357}.
CC -!- SUBCELLULAR LOCATION: Secreted {ECO:0000269|PubMed:15226357}. Host
CC cytoplasm, host cytosol {ECO:0000269|PubMed:15226357,
CC ECO:0000269|PubMed:17101670}. Host endoplasmic reticulum lumen
CC {ECO:0000305|PubMed:17024087, ECO:0000305|PubMed:18005237,
CC ECO:0000305|PubMed:23921389}. Note=Colocalizes with host BiP/HSPA5 in
CC the endoplasmic reticulum of Vero cells, its activity on BiP/HSPA5 is
CC blocked by pretreatment with brefeldin A, which disrupts the Golgi
CC apparatus and inhibits retrograde transport from the cell surface to
CC the Golgi (PubMed:17024087, PubMed:18005237). Using different
CC inhibitors it has been shown to be actively internalized by membrane-
CC bound vesicles and undergoes clathrin-dependent retrograde transport,
CC via early endosomes and the Golgi network, to the endoplasmic reticulum
CC (PubMed:18042253). Trafficking is similar in Vero cells, human HeLa
CC cells and murine N2A cells (PubMed:18042253).
CC {ECO:0000269|PubMed:15226357, ECO:0000269|PubMed:17024087,
CC ECO:0000269|PubMed:18005237, ECO:0000269|PubMed:18042253}.
CC -!- INDUCTION: Part of the subA-subB operon (PubMed:15226357).
CC {ECO:0000269|PubMed:15226357}.
CC -!- DOMAIN: The catalytic charge relay system lies at the bottom of a deep
CC cleft which probably accounts for its substrate specificity
CC (PubMed:17024087, PubMed:23921389). The C-terminal A2 domain
CC (approximately residues 322-347) penetrates into the central pore of
CC the SubB pentamer, making different contacts with each SubB subunit
CC (PubMed:23921389). {ECO:0000269|PubMed:17024087,
CC ECO:0000269|PubMed:23921389}.
CC -!- DISRUPTION PHENOTYPE: No longer toxic to CHO cells when deletion
CC construct is coexpressed with subB (PubMed:15226357).
CC {ECO:0000269|PubMed:15226357}.
CC -!- BIOTECHNOLOGY: Injection of the inactive mutant protein (Ala instead of
CC ser at position 272) in mice has no deleterious effects and elicits an
CC antibody response, suggesting it could be used as a vaccine
CC (PubMed:15226357). {ECO:0000269|PubMed:15226357}.
CC -!- MISCELLANEOUS: The E.coli strain this operon was isolated from causes
CC hemolytic uremic syndrome (HUS) and also encodes Stx, a Shigella-type
CC toxin. {ECO:0000303|PubMed:15226357}.
CC -!- SIMILARITY: Belongs to the peptidase S8 family. {ECO:0000305}.
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DR EMBL; AF399919; AAT68785.1; -; Genomic_DNA.
DR PDB; 2IY9; X-ray; 1.80 A; A=1-347.
DR PDB; 4BWG; X-ray; 2.60 A; A/G=1-347.
DR PDBsum; 2IY9; -.
DR PDBsum; 4BWG; -.
DR AlphaFoldDB; Q6EZC2; -.
DR SMR; Q6EZC2; -.
DR DIP; DIP-60412N; -.
DR IntAct; Q6EZC2; 1.
DR MEROPS; S08.121; -.
DR EvolutionaryTrace; Q6EZC2; -.
DR GO; GO:0005576; C:extracellular region; IEA:UniProtKB-SubCell.
DR GO; GO:0044164; C:host cell cytosol; IEA:UniProtKB-SubCell.
DR GO; GO:0044166; C:host cell endoplasmic reticulum lumen; IMP:UniProtKB.
DR GO; GO:0004252; F:serine-type endopeptidase activity; IMP:UniProtKB.
DR GO; GO:0090729; F:toxin activity; IEA:UniProtKB-KW.
DR GO; GO:0006508; P:proteolysis; IEA:UniProtKB-KW.
DR Gene3D; 3.40.50.200; -; 1.
DR InterPro; IPR000209; Peptidase_S8/S53_dom.
DR InterPro; IPR036852; Peptidase_S8/S53_dom_sf.
DR Pfam; PF00082; Peptidase_S8; 1.
DR SUPFAM; SSF52743; SSF52743; 1.
DR PROSITE; PS00014; ER_TARGET; 1.
DR PROSITE; PS51892; SUBTILASE; 1.
PE 1: Evidence at protein level;
KW 3D-structure; Direct protein sequencing; Disulfide bond; Host cytoplasm;
KW Host endoplasmic reticulum; Hydrolase; Plasmid; Protease; Secreted;
KW Serine protease; Signal; Toxin; Virulence.
FT SIGNAL 1..21
FT /evidence="ECO:0000255, ECO:0000305|PubMed:15226357"
FT CHAIN 22..347
FT /note="Subtilase cytotoxin subunit A"
FT /evidence="ECO:0000255"
FT /id="PRO_5004273014"
FT DOMAIN 24..327
FT /note="Peptidase S8"
FT /evidence="ECO:0000255|PROSITE-ProRule:PRU01240"
FT REGION 322..347
FT /note="A2 domain"
FT /evidence="ECO:0000269|PubMed:23921389"
FT MOTIF 344..347
FT /note="Prevents secretion from ER"
FT /evidence="ECO:0000255"
FT ACT_SITE 52
FT /note="Charge relay system"
FT /evidence="ECO:0000255|PROSITE-ProRule:PRU01240,
FT ECO:0000303|PubMed:15226357"
FT ACT_SITE 89
FT /note="Charge relay system"
FT /evidence="ECO:0000255|PROSITE-ProRule:PRU01240,
FT ECO:0000303|PubMed:15226357"
FT ACT_SITE 272
FT /note="Charge relay system"
FT /evidence="ECO:0000255|PROSITE-ProRule:PRU01240,
FT ECO:0000303|PubMed:15226357"
FT DISULFID 288..331
FT /evidence="ECO:0000269|PubMed:23921389,
FT ECO:0007744|PDB:2IY9, ECO:0007744|PDB:4BWG"
FT VARIANT 263
FT /note="A -> G (in strain: O29)"
FT /evidence="ECO:0000269|PubMed:17101670"
FT MUTAGEN 272
FT /note="S->A: Reduces cytotoxicity on Vero cells by more
FT than 99.9%, no toxic effect on mice when injected as
FT purified SubAB5. Does not cleave host BiP/HSPA5 in Vero
FT cells nor purified BiP/HSPA5, does not induce the unfolded
FT protein response in mice. No longer inhibits protein
FT synthesis in Vero cells. Does not induce apoptosis in Vero
FT cells."
FT /evidence="ECO:0000269|PubMed:15226357,
FT ECO:0000269|PubMed:17024087, ECO:0000269|PubMed:18005237,
FT ECO:0000269|PubMed:18433465, ECO:0000269|PubMed:19380466"
FT HELIX 26..30
FT /evidence="ECO:0007829|PDB:2IY9"
FT HELIX 34..38
FT /evidence="ECO:0007829|PDB:2IY9"
FT STRAND 47..53
FT /evidence="ECO:0007829|PDB:2IY9"
FT HELIX 59..61
FT /evidence="ECO:0007829|PDB:2IY9"
FT STRAND 65..70
FT /evidence="ECO:0007829|PDB:2IY9"
FT STRAND 81..83
FT /evidence="ECO:0007829|PDB:2IY9"
FT HELIX 84..98
FT /evidence="ECO:0007829|PDB:2IY9"
FT STRAND 100..103
FT /evidence="ECO:0007829|PDB:2IY9"
FT STRAND 111..116
FT /evidence="ECO:0007829|PDB:2IY9"
FT STRAND 119..121
FT /evidence="ECO:0007829|PDB:2IY9"
FT HELIX 125..134
FT /evidence="ECO:0007829|PDB:2IY9"
FT STRAND 143..151
FT /evidence="ECO:0007829|PDB:2IY9"
FT HELIX 160..172
FT /evidence="ECO:0007829|PDB:2IY9"
FT STRAND 176..180
FT /evidence="ECO:0007829|PDB:2IY9"
FT TURN 188..190
FT /evidence="ECO:0007829|PDB:2IY9"
FT TURN 193..195
FT /evidence="ECO:0007829|PDB:2IY9"
FT TURN 199..201
FT /evidence="ECO:0007829|PDB:2IY9"
FT HELIX 207..211
FT /evidence="ECO:0007829|PDB:2IY9"
FT STRAND 215..221
FT /evidence="ECO:0007829|PDB:2IY9"
FT STRAND 236..238
FT /evidence="ECO:0007829|PDB:2IY9"
FT TURN 244..246
FT /evidence="ECO:0007829|PDB:2IY9"
FT STRAND 248..252
FT /evidence="ECO:0007829|PDB:2IY9"
FT STRAND 254..259
FT /evidence="ECO:0007829|PDB:2IY9"
FT STRAND 265..268
FT /evidence="ECO:0007829|PDB:2IY9"
FT HELIX 271..288
FT /evidence="ECO:0007829|PDB:2IY9"
FT HELIX 294..304
FT /evidence="ECO:0007829|PDB:2IY9"
FT STRAND 305..307
FT /evidence="ECO:0007829|PDB:2IY9"
FT HELIX 309..311
FT /evidence="ECO:0007829|PDB:2IY9"
FT TURN 312..314
FT /evidence="ECO:0007829|PDB:2IY9"
FT HELIX 316..318
FT /evidence="ECO:0007829|PDB:2IY9"
FT HELIX 323..331
FT /evidence="ECO:0007829|PDB:2IY9"
FT HELIX 335..339
FT /evidence="ECO:0007829|PDB:4BWG"
SQ SEQUENCE 347 AA; 37494 MW; 708D010B14790B40 CRC64;
MLKILWTYIL FLLFISASAR AEKPWYFDAI GLTETTMSLT DKNTPVVVSV VDSGVAFIGG
LSDSEFAKFS FTQDGSPFPV KKSEALYIHG TAMASLIASR YGIYGVYPHA LISSRRVIPD
GVQDSWIRAI ESIMSNVFLA PGEEKIINIS GGQKGVASAS VWTELLSRMG RNNDRLIVAA
VGNDGADIRK LSAQQRIWPA AYHPVSSVNK KQDPVIRVAA LAQYRKGETP VLHGGGITGS
RFGNNWVDIA APGQNITFLR PDAKTGTGSG TSEATAIVSG VLAAMTSCNP RATATELKRT
LLESADKYPS LVDKVTEGRV LNAEKAISMF CKKNYIPVRQ GRMSEEL
