Escherichia coli K-12 substr. MG1655 Protein: ClpXP protease specificity-enhancing factor

Gene: sspB Accession Numbers: EG10978 (EcoCyc), b3228, ECK3217

Regulation Summary Diagram: ?

Regulation summary diagram for sspB

Subunit composition of ClpXP protease specificity-enhancing factor = [SspB]2

The SspB protein is a specificity-enhancing factor for the ClpXP protease [Levchenko00]. When protein synthesis is stalled, incomplete proteins that are produced are tagged with the small SsrA peptide. The ribosome-associated SspB protein binds to the SsrA tag and enhances degradation of the tagged peptide by the ClpXP protease [Levchenko00].

The SspB protein forms a homodimer with two independent binding sites for SsrA-tagged proteins [Wah02]. It also binds to ClpX and stimulates its ATPase activity [Wah02]. The dimerization and SsrA binding domain resides in the amino terminal 110-120 residues of SspB, while the C-terminal 40-50 residues are required for association with ClpXP and stimulation of its ATPase activity [Wah03]. Efficient ClpX hexamer binding and substrate delivery requires both C-terminal domains of the SspB dimer [Bolon04]. Interactions between binding of SspB and ClpX to the SsrA tag have been described [Hersch04].

Protein degradation substrates regulated by SspB have been identified and include RseA, which contains an SspB binding site that is unrelated to the SsrA tag sequence [Flynn04]. Degradation of the RseA cytoplasmic fragment is the last step in the proteolytic cascade leading to the induction of the sigma E extracytoplasmic stress regulon.

Crystal structures of SspB alone and in complex with the SsrA peptide tag have been determined at 2.2 and 2.9 A resolution, respectively [Song03], and a crystal structure of SspB in a complex with its recognition peptide in RseA has been determined at 1.8 A resolution [Levchenko05]. The crystal structures reveal diversity in the recognition of different target proteins.

The level of SspB protein remains constant throughout the transition from exponential growth to early stationary phase [Farrell05].

ssp: "stringent starvation protein"

Review: [Ades04]

Gene Citations: [Mason88, Williams91, Williams94, Kaczanowska04]

Locations: cytosol, ribosome

Map Position: [3,374,301 <- 3,374,798] (72.73 centisomes, 262°)
Length: 498 bp / 165 aa

Molecular Weight of Polypeptide: 18.262 kD (from nucleotide sequence)

Unification Links: ASAP:ABE-0010591 , CGSC:33594 , DIP:DIP-29563N , DisProt:DP00194 , EchoBASE:EB0971 , EcoGene:EG10978 , EcoliWiki:b3228 , Mint:MINT-1226453 , ModBase:P0AFZ3 , OU-Microarray:b3228 , PortEco:sspB , PR:PRO_000023993 , Pride:P0AFZ3 , Protein Model Portal:P0AFZ3 , RefSeq:NP_417695 , RegulonDB:EG10978 , SMR:P0AFZ3 , String:511145.b3228 , Swiss-Model:P0AFZ3 , UniProt:P0AFZ3

Relationship Links: InterPro:IN-FAMILY:IPR007481 , PDB:Structure:1OX8 , PDB:Structure:1OX9 , PDB:Structure:1YFN , PDB:Structure:2DS8 , Pfam:IN-FAMILY:PF04386

Gene-Reaction Schematic: ?

Gene-Reaction Schematic

GO Terms:

Biological Process: GO:0045732 - positive regulation of protein catabolic process Inferred from experiment [Levchenko00]
Molecular Function: GO:0005515 - protein binding Inferred from experiment [Wah02]
Cellular Component: GO:0005829 - cytosol Inferred from experiment [Ishihama08, LopezCampistrou05]
GO:0005840 - ribosome Inferred from experiment [Levchenko00]

MultiFun Terms: cell processes adaptations starvation

Essentiality data for sspB knockouts: ?

Growth Medium Growth? T (°C) O2 pH Osm/L Growth Observations
LB enriched Yes 37 Aerobic 6.95   Yes [Gerdes03, Comment 1]
LB Lennox Yes 37 Aerobic 7   Yes [Baba06, Comment 2]
M9 medium with 1% glycerol Yes 37 Aerobic 7.2 0.35 Yes [Joyce06, Comment 3]
MOPS medium with 0.4% glucose Yes 37 Aerobic 7.2 0.22 Yes [Baba06, Comment 2]

Sequence Features

Protein sequence of SspB with features indicated

Feature Class Location Citations Comment
Mutagenesis-Variant 52
[Levchenko05, UniProt14]
[Levchenko05, UniProt14]
V → I: 2.5-fold reduction in binding of RseA, no effect on ssrA-tag binding.
V → D, M or Q: No folded protein.
Mutagenesis-Variant 59
[Levchenko05, UniProt14]
UniProt: 10-fold reduction in binding of ssrA tag and RseA.
Protein-Segment 73 -> 95
UniProt: Binds SspB; Sequence Annotation Type: region of interest.
Mutagenesis-Variant 74
[Levchenko05, UniProt14]
UniProt: Severe reduction in binding of ssrA tag (40-fold) and RseA (100-fold).
Protein-Segment 128 -> 165
UniProt: Required for interaction with ClpX, not required for ssrA tag recognition or dimerization; Sequence Annotation Type: region of interest.

Gene Local Context (not to scale): ?

Gene local context diagram

Transcription Unit:

Transcription-unit diagram


10/20/97 Gene b3228 from Blattner lab Genbank (v. M52) entry merged into EcoCyc gene EG10978; confirmed by SwissProt match.


Ades04: Ades SE (2004). "Proteolysis: Adaptor, adaptor, catch me a catch." Curr Biol 14(21);R924-6. PMID: 15530384

Baba06: Baba T, Ara T, Hasegawa M, Takai Y, Okumura Y, Baba M, Datsenko KA, Tomita M, Wanner BL, Mori H (2006). "Construction of Escherichia coli K-12 in-frame, single-gene knockout mutants: the Keio collection." Mol Syst Biol 2;2006.0008. PMID: 16738554

Bolon04: Bolon DN, Wah DA, Hersch GL, Baker TA, Sauer RT (2004). "Bivalent tethering of SspB to ClpXP is required for efficient substrate delivery: a protein-design study." Mol Cell 13(3);443-9. PMID: 14967151

Farrell05: Farrell CM, Grossman AD, Sauer RT (2005). "Cytoplasmic degradation of ssrA-tagged proteins." Mol Microbiol 57(6);1750-61. PMID: 16135238

Flynn04: Flynn JM, Levchenko I, Sauer RT, Baker TA (2004). "Modulating substrate choice: the SspB adaptor delivers a regulator of the extracytoplasmic-stress response to the AAA+ protease ClpXP for degradation." Genes Dev 18(18);2292-301. PMID: 15371343

Gerdes03: Gerdes SY, Scholle MD, Campbell JW, Balazsi G, Ravasz E, Daugherty MD, Somera AL, Kyrpides NC, Anderson I, Gelfand MS, Bhattacharya A, Kapatral V, D'Souza M, Baev MV, Grechkin Y, Mseeh F, Fonstein MY, Overbeek R, Barabasi AL, Oltvai ZN, Osterman AL (2003). "Experimental determination and system level analysis of essential genes in Escherichia coli MG1655." J Bacteriol 185(19);5673-84. PMID: 13129938

Hersch04: Hersch GL, Baker TA, Sauer RT (2004). "SspB delivery of substrates for ClpXP proteolysis probed by the design of improved degradation tags." Proc Natl Acad Sci U S A 101(33);12136-41. PMID: 15297609

Ishihama08: Ishihama Y, Schmidt T, Rappsilber J, Mann M, Hartl FU, Kerner MJ, Frishman D (2008). "Protein abundance profiling of the Escherichia coli cytosol." BMC Genomics 9;102. PMID: 18304323

Joyce06: Joyce AR, Reed JL, White A, Edwards R, Osterman A, Baba T, Mori H, Lesely SA, Palsson BO, Agarwalla S (2006). "Experimental and computational assessment of conditionally essential genes in Escherichia coli." J Bacteriol 188(23);8259-71. PMID: 17012394

Kaczanowska04: Kaczanowska M, Ryden-Aulin M (2004). "Temperature sensitivity caused by mutant release factor 1 is suppressed by mutations that affect 16S rRNA maturation." J Bacteriol 186(10);3046-55. PMID: 15126466

Levchenko00: Levchenko I, Seidel M, Sauer RT, Baker TA (2000). "A specificity-enhancing factor for the ClpXP degradation machine." Science 289(5488);2354-6. PMID: 11009422

Levchenko05: Levchenko I, Grant RA, Flynn JM, Sauer RT, Baker TA (2005). "Versatile modes of peptide recognition by the AAA+ adaptor protein SspB." Nat Struct Mol Biol 12(6);520-5. PMID: 15880122

LopezCampistrou05: Lopez-Campistrous A, Semchuk P, Burke L, Palmer-Stone T, Brokx SJ, Broderick G, Bottorff D, Bolch S, Weiner JH, Ellison MJ (2005). "Localization, annotation, and comparison of the Escherichia coli K-12 proteome under two states of growth." Mol Cell Proteomics 4(8);1205-9. PMID: 15911532

Mason88: Mason JM, Hackett RH, Setlow P (1988). "Regulation of expression of genes coding for small, acid-soluble proteins of Bacillus subtilis spores: studies using lacZ gene fusions." J Bacteriol 1988;170(1);239-44. PMID: 3121585

Song03: Song HK, Eck MJ (2003). "Structural basis of degradation signal recognition by SspB, a specificity-enhancing factor for the ClpXP proteolytic machine." Mol Cell 12(1);75-86. PMID: 12887894

UniProt14: UniProt Consortium (2014). "UniProt version 2014-08 released on 2014-08-01 00:00:00." Database.

Wah02: Wah DA, Levchenko I, Baker TA, Sauer RT (2002). "Characterization of a Specificity Factor for an AAA+ ATPase. Assembly of SspB Dimers with ssrA-Tagged Proteins and the ClpX Hexamer." Chem Biol 9(11);1237-45. PMID: 12445774

Wah03: Wah DA, Levchenko I, Rieckhof GE, Bolon DN, Baker TA, Sauer RT (2003). "Flexible linkers leash the substrate binding domain of SspB to a peptide module that stabilizes delivery complexes with the AAA+ ClpXP protease." Mol Cell 12(2);355-63. PMID: 14536075

Williams91: Williams MD, Fuchs JA, Flickinger MC (1991). "Null mutation in the stringent starvation protein of Escherichia coli disrupts lytic development of bacteriophage P1." Gene 1991;109(1);21-30. PMID: 1721886

Williams94: Williams MD, Ouyang TX, Flickinger MC (1994). "Starvation-induced expression of SspA and SspB: the effects of a null mutation in sspA on Escherichia coli protein synthesis and survival during growth and prolonged starvation." Mol Microbiol 1994;11(6);1029-43. PMID: 8022275

Other References Related to Gene Regulation

Serizawa87: Serizawa H, Fukuda R (1987). "Structure of the gene for the stringent starvation protein of Escherichia coli." Nucleic Acids Res 1987;15(3);1153-63. PMID: 3029697

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Please cite the following article in publications resulting from the use of EcoCyc: Nucleic Acids Research 41:D605-12 2013
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