ID SPOT_ECOLI Reviewed; 702 AA. AC P0AG24; P17580; Q2M7W3; DT 20-DEC-2005, integrated into UniProtKB/Swiss-Prot. DT 20-DEC-2005, sequence version 1. DT 27-MAR-2024, entry version 145. DE RecName: Full=Bifunctional (p)ppGpp synthase/hydrolase SpoT; DE Includes: DE RecName: Full=GTP pyrophosphokinase; DE EC=2.7.6.5; DE AltName: Full=(p)ppGpp synthase; DE AltName: Full=ATP:GTP 3'-pyrophosphotransferase; DE AltName: Full=Stringent response-like protein; DE AltName: Full=ppGpp synthase II; DE Includes: DE RecName: Full=Guanosine-3',5'-bis(diphosphate) 3'-pyrophosphohydrolase; DE EC=3.1.7.2; DE AltName: Full=Penta-phosphate guanosine-3'-pyrophosphohydrolase; DE Short=(ppGpp)ase; GN Name=spoT; OrderedLocusNames=b3650, JW3625; OS Escherichia coli (strain K12). OC Bacteria; Pseudomonadota; Gammaproteobacteria; Enterobacterales; OC Enterobacteriaceae; Escherichia. OX NCBI_TaxID=83333; RN [1] RP NUCLEOTIDE SEQUENCE [GENOMIC DNA]. RX PubMed=2549050; DOI=10.1016/s0021-9258(18)63813-x; RA Sarubbi E., Rudd K.E., Xiao H., Ikehara K., Kalman M., Cashel M.; RT "Characterization of the spoT gene of Escherichia coli."; RL J. Biol. Chem. 264:15074-15082(1989). RN [2] RP NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA]. RC STRAIN=K12 / MG1655 / ATCC 47076; RX PubMed=7686882; DOI=10.1006/geno.1993.1230; RA Burland V.D., Plunkett G. III, Daniels D.L., Blattner F.R.; RT "DNA sequence and analysis of 136 kilobases of the Escherichia coli genome: RT organizational symmetry around the origin of replication."; RL Genomics 16:551-561(1993). RN [3] RP NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA]. RC STRAIN=K12 / MG1655 / ATCC 47076; RX PubMed=9278503; DOI=10.1126/science.277.5331.1453; RA Blattner F.R., Plunkett G. III, Bloch C.A., Perna N.T., Burland V., RA Riley M., Collado-Vides J., Glasner J.D., Rode C.K., Mayhew G.F., RA Gregor J., Davis N.W., Kirkpatrick H.A., Goeden M.A., Rose D.J., Mau B., RA Shao Y.; RT "The complete genome sequence of Escherichia coli K-12."; RL Science 277:1453-1462(1997). RN [4] RP NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA]. RC STRAIN=K12 / W3110 / ATCC 27325 / DSM 5911; RX PubMed=16738553; DOI=10.1038/msb4100049; RA Hayashi K., Morooka N., Yamamoto Y., Fujita K., Isono K., Choi S., RA Ohtsubo E., Baba T., Wanner B.L., Mori H., Horiuchi T.; RT "Highly accurate genome sequences of Escherichia coli K-12 strains MG1655 RT and W3110."; RL Mol. Syst. Biol. 2:E1-E5(2006). RN [5] RP PROBABLE (P)PPGPP SYNTHESIS ACTIVITY. RX PubMed=2005134; DOI=10.1016/s0021-9258(19)67694-5; RA Xiao H., Kalman M., Ikehara K., Zemel S., Glaser G., Cashel M.; RT "Residual guanosine 3',5'-bispyrophosphate synthetic activity of relA null RT mutants can be eliminated by spoT null mutations."; RL J. Biol. Chem. 266:5980-5990(1991). RN [6] RP PPGPP SYNTHASE ACTIVITY, AND MUTAGENESIS OF ASP-293. RC STRAIN=CF8295; RX PubMed=12596879; DOI=10.1271/bbb.66.2735; RA Fujita C., Maeda M., Fujii T., Iwamoto R., Ikehara K.; RT "Identification of an indispensable amino acid for ppGpp synthesis of RT Escherichia coli SpoT protein."; RL Biosci. Biotechnol. Biochem. 66:2735-2738(2002). RN [7] RP FUNCTION IN PERSISTENCE, AND DISRUPTION PHENOTYPE. RC STRAIN=K12 / MG1655 / ATCC 47076; RX PubMed=14622409; DOI=10.1046/j.1365-2958.2003.03779.x; RA Korch S.B., Henderson T.A., Hill T.M.; RT "Characterization of the hipA7 allele of Escherichia coli and evidence that RT high persistence is governed by (p)ppGpp synthesis."; RL Mol. Microbiol. 50:1199-1213(2003). RN [8] RP INTERACTION WITH OBGE/CGTA. RC STRAIN=K12 / W3110 / ATCC 27325 / DSM 5911; RX PubMed=15292126; DOI=10.1128/jb.186.16.5249-5257.2004; RA Wout P., Pu K., Sullivan S.M., Reese V., Zhou S., Lin B., Maddock J.R.; RT "The Escherichia coli GTPase CgtAE cofractionates with the 50S ribosomal RT subunit and interacts with SpoT, a ppGpp synthetase/hydrolase."; RL J. Bacteriol. 186:5249-5257(2004). RN [9] RP SUBCELLULAR LOCATION, LARGE RIBOSOMAL SUBUNIT ASSOCIATION, AND RP CHARACTERIZATION UNDER AMINO ACID OR CARBON STARVATION. RC STRAIN=K12; RX PubMed=17616600; DOI=10.1128/jb.00315-07; RA Jiang M., Sullivan S.M., Wout P.K., Maddock J.R.; RT "G-protein control of the ribosome-associated stress response protein RT SpoT."; RL J. Bacteriol. 189:6140-6147(2007). RN [10] RP FUNCTION IN BIOFILM FORMATION, MUTAGENESIS OF ASP-73 AND ASP-259, AND RP DISRUPTION PHENOTYPE. RC STRAIN=K12 / MG1655 / AB400; RX PubMed=19460094; DOI=10.1111/j.1365-2958.2009.06739.x; RA Boehm A., Steiner S., Zaehringer F., Casanova A., Hamburger F., Ritz D., RA Keck W., Ackermann M., Schirmer T., Jenal U.; RT "Second messenger signalling governs Escherichia coli biofilm induction RT upon ribosomal stress."; RL Mol. Microbiol. 72:1500-1516(2009). RN [11] RP FUNCTION, AND DISRUPTION PHENOTYPE. RC STRAIN=K12 / CF7789, and K12 / MG1655 / ATCC 47076; RX PubMed=21488981; DOI=10.1111/j.1365-2958.2011.07663.x; RA Edwards A.N., Patterson-Fortin L.M., Vakulskas C.A., Mercante J.W., RA Potrykus K., Vinella D., Camacho M.I., Fields J.A., Thompson S.A., RA Georgellis D., Cashel M., Babitzke P., Romeo T.; RT "Circuitry linking the Csr and stringent response global regulatory RT systems."; RL Mol. Microbiol. 80:1561-1580(2011). RN [12] RP FUNCTION IN PERSISTENCE, AND DISRUPTION PHENOTYPE. RC STRAIN=K12 / BW25113; RX PubMed=26051177; DOI=10.1016/j.molcel.2015.05.011; RA Verstraeten N., Knapen W.J., Kint C.I., Liebens V., Van den Bergh B., RA Dewachter L., Michiels J.E., Fu Q., David C.C., Fierro A.C., Marchal K., RA Beirlant J., Versees W., Hofkens J., Jansen M., Fauvart M., Michiels J.; RT "Obg and membrane depolarization are part of a microbial bet-hedging RT strategy that leads to antibiotic tolerance."; RL Mol. Cell 59:9-21(2015). CC -!- FUNCTION: In eubacteria ppGpp (guanosine 3'-diphosphate 5'-diphosphate) CC is a mediator of the stringent response which coordinates a variety of CC cellular activities in response to changes in nutritional abundance. CC This enzyme catalyzes both the synthesis and degradation of ppGpp. The CC second messengers ppGpp and c-di-GMP together control biofilm formation CC in response to translational stress; ppGpp represses biofilm formation CC while c-di-GMP induces it. ppGpp activates transcription of CsrA- CC antagonistic small RNAs CsrB and CsrC, which down-regulate CsrA's CC action on translation during the stringent response (PubMed:21488981). CC {ECO:0000269|PubMed:14622409, ECO:0000269|PubMed:19460094, CC ECO:0000269|PubMed:21488981}. CC -!- CATALYTIC ACTIVITY: CC Reaction=ATP + GTP = AMP + guanosine 3'-diphosphate 5'-triphosphate; CC Xref=Rhea:RHEA:22088, ChEBI:CHEBI:30616, ChEBI:CHEBI:37565, CC ChEBI:CHEBI:142410, ChEBI:CHEBI:456215; EC=2.7.6.5; CC -!- CATALYTIC ACTIVITY: CC Reaction=guanosine 3',5'-bis(diphosphate) + H2O = diphosphate + GDP + CC H(+); Xref=Rhea:RHEA:14253, ChEBI:CHEBI:15377, ChEBI:CHEBI:15378, CC ChEBI:CHEBI:33019, ChEBI:CHEBI:58189, ChEBI:CHEBI:77828; EC=3.1.7.2; CC -!- COFACTOR: CC Name=Mn(2+); Xref=ChEBI:CHEBI:29035; CC -!- PATHWAY: Purine metabolism; ppGpp biosynthesis; ppGpp from GDP: step CC 1/1. CC -!- PATHWAY: Purine metabolism; ppGpp biosynthesis; ppGpp from GTP: step CC 1/2. CC -!- SUBUNIT: Interacts with ObgE/CgtA (AC P42641); this association is not CC required for pre-50S ribosome binding. Under both amino acid and carbon CC starvation conditions about half of the protein dissociates from the CC ribosome. {ECO:0000269|PubMed:15292126}. CC -!- INTERACTION: CC P0AG24; P0A6A8: acpP; NbExp=6; IntAct=EBI-543228, EBI-542566; CC -!- SUBCELLULAR LOCATION: Cytoplasm {ECO:0000269|PubMed:17616600}. Note=Is CC associated with pre-50S ribosomal subunits in a salt-dependent manner. CC -!- DISRUPTION PHENOTYPE: In the presence of wild-type relA, spoT cannot be CC deleted because ppGpp accumulation is toxic. In the double relA/spoT CC deletion (a ppGpp0 mutant) there is a very large increase in biofilm CC formation (in a csrA-disrupted background) (PubMed:19460094). The CC ppGpp0 mutant makes decreased levels of CsrA and its inhibitory small CC RNAs (sRNA) CsrB and CsrC (PubMed:21488981). The double relA/spoT CC deletion obviates persister cell formation in a hipA7 mutant CC (PubMed:14622409, PubMed:26051177). {ECO:0000269|PubMed:14622409, CC ECO:0000269|PubMed:19460094, ECO:0000269|PubMed:21488981, CC ECO:0000269|PubMed:26051177}. CC -!- SIMILARITY: Belongs to the RelA/SpoT family. {ECO:0000305}. CC --------------------------------------------------------------------------- CC Copyrighted by the UniProt Consortium, see https://www.uniprot.org/terms CC Distributed under the Creative Commons Attribution (CC BY 4.0) License CC --------------------------------------------------------------------------- DR EMBL; M24503; AAB00160.1; -; Genomic_DNA. DR EMBL; L10328; AAA62003.1; -; Genomic_DNA. DR EMBL; U00096; AAC76674.1; -; Genomic_DNA. DR EMBL; AP009048; BAE77643.1; -; Genomic_DNA. DR PIR; B30374; SHECGD. DR RefSeq; NP_418107.1; NC_000913.3. DR RefSeq; WP_000280488.1; NZ_STEB01000024.1. DR AlphaFoldDB; P0AG24; -. DR SMR; P0AG24; -. DR BioGRID; 4259361; 494. DR DIP; DIP-29378N; -. DR IntAct; P0AG24; 61. DR STRING; 511145.b3650; -. DR jPOST; P0AG24; -. DR PaxDb; 511145-b3650; -. DR EnsemblBacteria; AAC76674; AAC76674; b3650. DR GeneID; 75202219; -. DR GeneID; 948159; -. DR KEGG; ecj:JW3625; -. DR KEGG; eco:b3650; -. DR PATRIC; fig|1411691.4.peg.3056; -. DR EchoBASE; EB0959; -. DR eggNOG; COG0317; Bacteria. DR HOGENOM; CLU_012300_3_0_6; -. DR InParanoid; P0AG24; -. DR OMA; PIDFAYS; -. DR PhylomeDB; P0AG24; -. DR BioCyc; EcoCyc:SPOT-MONOMER; -. DR BioCyc; MetaCyc:SPOT-MONOMER; -. DR BRENDA; 2.7.6.5; 2026. DR BRENDA; 3.1.7.2; 2026. DR UniPathway; UPA00908; UER00884. DR UniPathway; UPA00908; UER00886. DR PRO; PR:P0AG24; -. DR Proteomes; UP000000318; Chromosome. DR Proteomes; UP000000625; Chromosome. DR GO; GO:0005829; C:cytosol; IDA:EcoCyc. DR GO; GO:0008728; F:GTP diphosphokinase activity; IBA:GO_Central. DR GO; GO:0008893; F:guanosine-3',5'-bis(diphosphate) 3'-diphosphatase activity; IDA:EcoCyc. DR GO; GO:0016301; F:kinase activity; IEA:UniProtKB-KW. DR GO; GO:0015970; P:guanosine tetraphosphate biosynthetic process; IEA:UniProtKB-UniPathway. DR GO; GO:0015969; P:guanosine tetraphosphate metabolic process; IBA:GO_Central. DR GO; GO:0015949; P:nucleobase-containing small molecule interconversion; IMP:EcoCyc. DR GO; GO:0016310; P:phosphorylation; IEA:UniProtKB-KW. DR GO; GO:0042594; P:response to starvation; IBA:GO_Central. DR CDD; cd04876; ACT_RelA-SpoT; 1. DR CDD; cd00077; HDc; 1. DR CDD; cd05399; NT_Rel-Spo_like; 1. DR CDD; cd01668; TGS_RSH; 1. DR Gene3D; 3.10.20.30; -; 1. DR Gene3D; 3.30.70.260; -; 1. DR Gene3D; 3.30.460.10; Beta Polymerase, domain 2; 1. DR Gene3D; 1.10.3210.10; Hypothetical protein af1432; 1. DR InterPro; IPR045865; ACT-like_dom_sf. DR InterPro; IPR002912; ACT_dom. DR InterPro; IPR012675; Beta-grasp_dom_sf. DR InterPro; IPR003607; HD/PDEase_dom. DR InterPro; IPR006674; HD_domain. DR InterPro; IPR043519; NT_sf. DR InterPro; IPR004811; RelA/Spo_fam. DR InterPro; IPR045600; RelA/SpoT_AH_RIS. DR InterPro; IPR007685; RelA_SpoT. DR InterPro; IPR004095; TGS. DR InterPro; IPR012676; TGS-like. DR InterPro; IPR033655; TGS_RelA/SpoT. DR NCBIfam; TIGR00691; spoT_relA; 1. DR PANTHER; PTHR21262:SF31; BIFUNCTIONAL (P)PPGPP SYNTHASE_HYDROLASE SPOT; 1. DR PANTHER; PTHR21262; GUANOSINE-3',5'-BIS DIPHOSPHATE 3'-PYROPHOSPHOHYDROLASE; 1. DR Pfam; PF13291; ACT_4; 1. DR Pfam; PF13328; HD_4; 1. DR Pfam; PF19296; RelA_AH_RIS; 1. DR Pfam; PF04607; RelA_SpoT; 1. DR Pfam; PF02824; TGS; 1. DR SMART; SM00471; HDc; 1. DR SMART; SM00954; RelA_SpoT; 1. DR SUPFAM; SSF55021; ACT-like; 1. DR SUPFAM; SSF109604; HD-domain/PDEase-like; 1. DR SUPFAM; SSF81301; Nucleotidyltransferase; 1. DR SUPFAM; SSF81271; TGS-like; 1. DR PROSITE; PS51671; ACT; 1. DR PROSITE; PS51831; HD; 1. DR PROSITE; PS51880; TGS; 1. PE 1: Evidence at protein level; KW Cytoplasm; Hydrolase; Kinase; Manganese; Reference proteome; Transferase. FT CHAIN 1..702 FT /note="Bifunctional (p)ppGpp synthase/hydrolase SpoT" FT /id="PRO_0000166569" FT DOMAIN 45..144 FT /note="HD" FT /evidence="ECO:0000255|PROSITE-ProRule:PRU01175" FT DOMAIN 386..447 FT /note="TGS" FT /evidence="ECO:0000255|PROSITE-ProRule:PRU01228" FT DOMAIN 628..702 FT /note="ACT" FT /evidence="ECO:0000255|PROSITE-ProRule:PRU01007" FT MUTAGEN 73 FT /note="D->N: Obviates hydrolysis of ppGpp, moderately FT decreases biofilm formation, loss of biofilm induction in FT response to translation inhibitors." FT /evidence="ECO:0000269|PubMed:19460094" FT MUTAGEN 259 FT /note="D->N: Obviates synthesis of ppGpp, strongly FT increases biofilm formation." FT /evidence="ECO:0000269|PubMed:19460094" FT MUTAGEN 293 FT /note="D->A: Unable to restore (p)ppGpp synthesis nor FT complement a relA/spoT double disruption." FT /evidence="ECO:0000269|PubMed:12596879" FT MUTAGEN 294 FT /note="Y->I: Synthesizes (p)ppGpp, complements a relA/spoT FT double disruption." SQ SEQUENCE 702 AA; 79342 MW; A85F70F57D082EE8 CRC64; MYLFESLNQL IQTYLPEDQI KRLRQAYLVA RDAHEGQTRS SGEPYITHPV AVACILAEMK LDYETLMAAL LHDVIEDTPA TYQDMEQLFG KSVAELVEGV SKLDKLKFRD KKEAQAENFR KMIMAMVQDI RVILIKLADR THNMRTLGSL RPDKRRRIAR ETLEIYSPLA HRLGIHHIKT ELEELGFEAL YPNRYRVIKE VVKAARGNRK EMIQKILSEI EGRLQEAGIP CRVSGREKHL YSIYCKMVLK EQRFHSIMDI YAFRVIVNDS DTCYRVLGQM HSLYKPRPGR VKDYIAIPKA NGYQSLHTSM IGPHGVPVEV QIRTEDMDQM AEMGVAAHWA YKEHGETSTT AQIRAQRWMQ SLLELQQSAG SSFEFIESVK SDLFPDEIYV FTPEGRIVEL PAGATPVDFA YAVHTDIGHA CVGARVDRQP YPLSQPLTSG QTVEIITAPG ARPNAAWLNF VVSSKARAKI RQLLKNLKRD DSVSLGRRLL NHALGGSRKL NEIPQENIQR ELDRMKLATL DDLLAEIGLG NAMSVVVAKN LQHGDASIPP ATQSHGHLPI KGADGVLITF AKCCRPIPGD PIIAHVSPGK GLVIHHESCR NIRGYQKEPE KFMAVEWDKE TAQEFITEIK VEMFNHQGAL ANLTAAINTT TSNIQSLNTE EKDGRVYSAF IRLTARDRVH LANIMRKIRV MPDVIKVTRN RN //