SpoT

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Figure 1: Predicted structure of bifunctional (p)ppGpp synthase/hydrolase SpoT. Dark blue areas represent very high model confidence (pLDDT > 90). Light blue areas represent moderate model confidence (90>pLDDT > 70). Yellow areas represent low model confidence (70>pLDDT > 50). Orange areas represent very low model confidence (pLDDT < 50). Bifunctional ppGpp synthase hydrolase SpoT.png
Figure 1: Predicted structure of bifunctional (p)ppGpp synthase/hydrolase SpoT. Dark blue areas represent very high model confidence (pLDDT > 90). Light blue areas represent moderate model confidence (90>pLDDT > 70). Yellow areas represent low model confidence (70>pLDDT > 50). Orange areas represent very low model confidence (pLDDT < 50).

Bifunctional (p)ppGpp synthase/hydrolase SpoT or SpoT is a regulatory enzyme in the RelA/SpoT Homologue (RSH) protein family that synthesizes and hydrolyzes (p)ppGpp to regulate the bacterial stringent response to environmental stressors. [3] SpoT is considered a "long" form RSH protein and is found in many bacteria and plant chloroplasts. [4] SpoT and its homologues have been studied in bacterial model organism E.coli for their role in the production and degradation of (p)ppGpp in the stringent response pathway.

Contents

Bifunctional (p)ppGpp synthase/hydrolase SpoT
Identifiers
Organism Escherichia coli (strain K12)
SymbolspoT
UniProt P0AG24
Search for
Structures Swiss-model
Domains InterPro

Role in Stringent Response Pathway

The stringent response regulated by SpoT, RelA, and their homologues can cause a bacterium to increase its persistence in stressful environments. [5] SpoT can act as both a hydrolase and a synthetase to (p)ppGpp alarmones in the stringent response pathway with Mn2+ as its cofactor. [6] When there are environmental stressors present, SpoT uses ATP and GDP to synthesize (p)ppGpp and catalyze the stringent response. [7] When stressors are removed and a stringent response is no longer necessary SpoT hydrolyzes (p)ppGpp, cleaving it into GTP and diphosphate. [7] Environmental stressors including but not limited to amino acid starvation, [3] carbon deficiencies, [8] phosphate deficiencies [9] and changes in temperature [4] have been documented to cause the gene encoding SpoT to activate.

The acyl carrier protein (ACP) binds to the TGS domain of SpoT; this binding is probably influenced by the ratio of unacylated ACP to acylated ACP in the cell. [10]

Role as a Hydrolase

SpoT mainly serves as a hydrolase in systems similar to E.coli. SpoT's hydrolase activity is Mn2+-dependent with a conserved His-Asp (HD) motif. [11] Phosphate starvation is sensed by SpoT hydrolase to elevate (p)ppGpp, which induces IraP, a RssB antiadaptor that antagonizes RssB activation of RpoS turnover, thereby inducing RpoS. [9]

SpoT in E. coli

In E. coli, the SpoT protein consists of 702 amino acids. [6] E.coli uses RelA and SpoT as its two main (p)ppGpp regulating enzymes. [12] When the gene for encoding RelA is nonfunctional, E. coli can still regulate (p)ppGpp through SpoT as it has both HD and SYNTH domains.

Figure 2: RelA/SpoT Homologue (RSH) protein family domains separated by hydrolase (HD) function and synthetase (SYNTH) function. Pone.0023479.g001.png
Figure 2: RelA/SpoT Homologue (RSH) protein family domains separated by hydrolase (HD) function and synthetase (SYNTH) function.

SpoT and RelA have many homologous variations, forming the RelA/SpoT Homologue (RSH) protein family. These homologues serve similar functions to SpoT and RelA in stringent responses. Protein domains observed in members of the RSH protein family are separated by hydrolase (HD) functionality and synthetase (SYNTH) functionality (see Figure 2). [4]

Related Research Articles

<span class="mw-page-title-main">Nucleoside-diphosphate kinase</span> Class of enzymes

Nucleoside-diphosphate kinases are enzymes that catalyze the exchange of terminal phosphate between different nucleoside diphosphates (NDP) and triphosphates (NTP) in a reversible manner to produce nucleotide triphosphates. Many NDP serve as acceptor while NTP are donors of phosphate group. The general reaction via ping-pong mechanism is as follows: XDP + YTP ←→ XTP + YDP. NDPK activities maintain an equilibrium between the concentrations of different nucleoside triphosphates such as, for example, when guanosine triphosphate (GTP) produced in the citric acid (Krebs) cycle is converted to adenosine triphosphate (ATP). Other activities include cell proliferation, differentiation and development, signal transduction, G protein-coupled receptor, endocytosis, and gene expression.

<span class="mw-page-title-main">Glutamine synthetase</span> Class of enzymes

Glutamine synthetase (GS) is an enzyme that plays an essential role in the metabolism of nitrogen by catalyzing the condensation of glutamate and ammonia to form glutamine:

The stringent response, also called stringent control, is a stress response of bacteria and plant chloroplasts in reaction to amino-acid starvation, fatty acid limitation, iron limitation, heat shock and other stress conditions. The stringent response is signaled by the alarmone (p)ppGpp, and modulates transcription of up to 1/3 of all genes in the cell. This in turn causes the cell to divert resources away from growth and division and toward amino acid synthesis in order to promote survival until nutrient conditions improve.

The gene rpoS encodes the sigma factor sigma-38, a 37.8 kD protein in Escherichia coli. Sigma factors are proteins that regulate transcription in bacteria. Sigma factors can be activated in response to different environmental conditions. rpoS is transcribed in late exponential phase, and RpoS is the primary regulator of stationary phase genes. RpoS is a central regulator of the general stress response and operates in both a retroactive and a proactive manner: it not only allows the cell to survive environmental challenges, but it also prepares the cell for subsequent stresses (cross-protection). The transcriptional regulator CsgD is central to biofilm formation, controlling the expression of the curli structural and export proteins, and the diguanylate cyclase, adrA, which indirectly activates cellulose production. The rpoS gene most likely originated in the gammaproteobacteria.

In molecular genetics, a regulon is a group of genes that are regulated as a unit, generally controlled by the same regulatory gene that expresses a protein acting as a repressor or activator. This terminology is generally, although not exclusively, used in reference to prokaryotes, whose genomes are often organized into operons; the genes contained within a regulon are usually organized into more than one operon at disparate locations on the chromosome. Applied to eukaryotes, the term refers to any group of non-contiguous genes controlled by the same regulatory gene.

<span class="mw-page-title-main">CTP synthetase</span> Enzyme

CTP synthase is an enzyme involved in pyrimidine biosynthesis that interconverts UTP and CTP.

<span class="mw-page-title-main">MicF RNA</span> Gene found in bacteria

The micF RNA is a non-coding RNA stress response gene found in Escherichia coli and related bacteria that post-transcriptionally controls expression of the outer membrane porin gene ompF. The micF gene encodes a non-translated 93 nucleotide antisense RNA that binds its target ompF mRNA and regulates ompF expression by inhibiting translation and inducing degradation of the message. In addition, other factors, such as the RNA chaperone protein StpA also play a role in this regulatory system. The expression of micF is controlled by both environmental and internal stress factors. Four transcriptional regulators are known to bind the micF promoter region and activate micF expression.

<span class="mw-page-title-main">SgrS RNA</span>

SgrS is a 227 nucleotide small RNA that is activated by SgrR in Escherichia coli during glucose-phosphate stress. The nature of glucose-phosphate stress is not fully understood, but is correlated with intracellular accumulation of glucose-6-phosphate. SgrS helps cells recover from glucose-phosphate stress by base pairing with ptsG mRNA and causing its degradation in an RNase E dependent manner. Base pairing between SgrS and ptsG mRNA also requires Hfq, an RNA chaperone frequently required by small RNAs that affect their targets through base pairing. The inability of cells expressing sgrS to create new glucose transporters leads to less glucose uptake and reduced levels of glucose-6-phosphate. SgrS is an unusual small RNA in that it also encodes a 43 amino acid functional polypeptide, SgrT, which helps cells recover from glucose-phosphate stress by preventing glucose uptake. The activity of SgrT does not affect the levels of ptsG mRNA of PtsG protein. It has been proposed that SgrT exerts its effects through regulation of the glucose transporter, PtsG.

<span class="mw-page-title-main">Hfq protein</span>

The Hfq protein encoded by the hfq gene was discovered in 1968 as an Escherichia coli host factor that was essential for replication of the bacteriophage Qβ. It is now clear that Hfq is an abundant bacterial RNA binding protein which has many important physiological roles that are usually mediated by interacting with Hfq binding sRNA.

<span class="mw-page-title-main">Hok/sok system</span>

The hok/sok system is a postsegregational killing mechanism employed by the R1 plasmid in Escherichia coli. It was the first type I toxin-antitoxin pair to be identified through characterisation of a plasmid-stabilising locus. It is a type I system because the toxin is neutralised by a complementary RNA, rather than a partnered protein.

<span class="mw-page-title-main">Guanosine pentaphosphate</span> Chemical compound

(p)ppGpp, guanosine pentaphosphate and tetraphosphate, also known as the "magic spot" nucleotides, are alarmones involved in the stringent response in bacteria that cause the inhibition of RNA synthesis when there is a shortage of amino acids. This inhibition by (p)ppGpp decreases translation in the cell, conserving amino acids present. Furthermore, ppGpp and pppGpp cause the up-regulation of many other genes involved in stress response such as the genes for amino acid uptake and biosynthesis.

The enzyme [acyl-carrier-protein] phosphodiesterase (EC 3.1.4.14) catalyzes the reaction

<span class="mw-page-title-main">Holo-(acyl-carrier-protein) synthase</span>

In enzymology and molecular biology, a holo-[acyl-carrier-protein] synthase is an enzyme that catalyzes the chemical reaction:

An alarmone is an intracellular signal molecule that is produced in bacteria, chloroplasts, and a slim minority of archaea reacting to harsh environmental factors. They regulate the gene expression at transcription level. Alarmones are produced in high concentrations when harsh environmental factors occur in bacteria and plants, such as lack of amino acids, to produce proteins. Stringent factors take uncharged tRNA and convert it to an alarmone. Guanosine-5'-triphosphate (GTP) is then converted to 5´-diphosphate 3´-diphosphate guanosine (ppGpp), the archetypical alarmone. ppGpp will bind to RNA polymerase β and β´ subunits, changing promoter preference. It will decrease transcription of rRNA and other genes but will increase transcription of genes involved in amino acid biosyntheses and metabolisms involved in famine.

Bacterial small RNAs (bsRNA) are small RNAs produced by bacteria; they are 50- to 500-nucleotide non-coding RNA molecules, highly structured and containing several stem-loops. Numerous sRNAs have been identified using both computational analysis and laboratory-based techniques such as Northern blotting, microarrays and RNA-Seq in a number of bacterial species including Escherichia coli, the model pathogen Salmonella, the nitrogen-fixing alphaproteobacterium Sinorhizobium meliloti, marine cyanobacteria, Francisella tularensis, Streptococcus pyogenes, the pathogen Staphylococcus aureus, and the plant pathogen Xanthomonas oryzae pathovar oryzae. Bacterial sRNAs affect how genes are expressed within bacterial cells via interaction with mRNA or protein, and thus can affect a variety of bacterial functions like metabolism, virulence, environmental stress response, and structure.

<span class="mw-page-title-main">Toxin-antitoxin system</span> Biological process

A toxin-antitoxin system consists of a "toxin" and a corresponding "antitoxin", usually encoded by closely linked genes. The toxin is usually a protein while the antitoxin can be a protein or an RNA. Toxin-antitoxin systems are widely distributed in prokaryotes, and organisms often have them in multiple copies. When these systems are contained on plasmids – transferable genetic elements – they ensure that only the daughter cells that inherit the plasmid survive after cell division. If the plasmid is absent in a daughter cell, the unstable antitoxin is degraded and the stable toxic protein kills the new cell; this is known as 'post-segregational killing' (PSK).

The gua operon is responsible for regulating the synthesis of guanosine mono phosphate (GMP), a purine nucleotide, from inosine monophosphate. It consists of two structural genes guaB (encodes for IMP dehydrogenase or and guaA apart from the promoter and operator region.

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A protein superfamily is the largest grouping (clade) of proteins for which common ancestry can be inferred. Usually this common ancestry is inferred from structural alignment and mechanistic similarity, even if no sequence similarity is evident. Sequence homology can then be deduced even if not apparent. Superfamilies typically contain several protein families which show sequence similarity within each family. The term protein clan is commonly used for protease and glycosyl hydrolases superfamilies based on the MEROPS and CAZy classification systems.

<span class="mw-page-title-main">Universal stress protein</span>

The universal stress protein (USP) domain is a superfamily of conserved genes which can be found in bacteria, archaea, fungi, protozoa and plants. Proteins containing the domain are induced by many environmental stressors such as nutrient starvation, drought, extreme temperatures, high salinity, and the presence of uncouplers, antibiotics and metals.

References

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Further reading