Trp Operon Vs Lac Operon

Tryptophan Operon of Escherichia coli

C. Yanofsky , in Brenner's Encyclopedia of Genetics (Second Edition), 2013

Abstract

The trp operon of E. coli contains five major structural genes encoding all seven poly peptide functional domains necessary for tryptophan biosynthesis from the common aromatic precursor, chorismate. Transcription of the trp operon is highly regulated. Initiation of transcription at the trp promoter is regulated by the tryptophan-activated trp repressor. The repressor can bind at multiple operator sites located in the promoter region. Transcription of the structural genes of the trp operon also is regulated by transcription attenuation, in response to the accumulation of uncharged tRNA^Trp. When this uncharged tRNA accumulates, it leads to ribosome stalling during attempted translation of the leader peptide coding region. This leads to the formation of an RNA antiterminator structure that prevents the RNA terminator structure from forming. Absence of the terminator structure allows polymerase to go on transcription into the structural genes of the operon. When there are adequate levels of charged tRNA^Trp in the cell, the ribosome translating the leader peptide coding region completes leader peptide synthesis, assuasive the RNA terminator structure to form, and finish transcription.

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Cistron Expression: Transcription of the Genetic Code

Chang-Hui Shen , in Diagnostic Molecular Biology, 2019

Transcription Attenuation

The trp operon of East. coli codes for the enzymes that the bacterium needs to make the amino acid tryptophan. Like the lac operon, the trp operon is a negative command mechanism. The lac operon responds to an inducer that causes the repressor to dissociate from the operator, derepressing the operon. The trp operon responds to a repressor protein that binds to 2 molecules of tryptophan. When the tryptophan is plentiful, this repressor-tryptophan complex binds to the trp operator. This binding prevents the binding of RNA polymerase, then the operon is not transcribed (Fig. 3.xx). On the other hand, when tryptophan levels are reduced, the repressor will non bind the operator, then the operon is transcribed. This is an example of a system that is repressible and under negative regulation.

In add-on to the standard negative regulation, the trp operon is regulated by another mechanism of control called transcription attenuation. This mechanism operates by causing premature termination of transcription of the operon when tryptophan is abundant. Every bit shown in Fig. 3.20, there are two loci, the trp leader and the trp attenuator, in between the operator and the gene trpE. Secondary structures formed in the mRNA of the leader sequence are responsible for this premature termination. The formation of such secondary structures comes from the transcription stop signals—an inverted repeat and a string of 8 A-T pairs in the attenuator. When tryptophan is deficient, the operon is translated usually. When it is plentiful, transcription is terminated prematurely after the leader sequences have been transcribed. Thus, attenuation imposes an extra level of command on an operon, over and in a higher place the repressor-operator organization.

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Regulation of Cistron Expression

Northward.V. Bhagavan , Chung-Eun Ha , in Essentials of Medical Biochemistry, 2011

Tryptophan (Trp) Operon

The tryptophan operon is responsible for the product of the amino acid tryptophan, whose synthesis occurs in five steps, each requiring a particular enzyme. In East. coli, these enzymes are translated from a unmarried polycistronic mRNA. Adjacent to the enzyme coding sequences in the DNA are a promoter, an operator, and two regions called the leader and the attenuator (Figure 24-three). The leader and attenuator sequences are transcribed. Another factor (trpR) encoding a repressor is located some altitude from this gene cluster.

Regulation of the trp operon is adamant by the concentration of tryptophan; when adequate tryptophan is nowadays in the growth medium, there is no need for tryptophan biosynthesis. Transcription is turned off when a loftier concentration of tryptophan is present, and is turned on when tryptophan is absent. The regulatory bespeak is the concentration of tryptophan itself. In contrast to lactose, tryptophan is active in repression rather than induction.

The trp operon has two levels of regulation: an on-off mechanism and a modulation system. The protein production of the trpR gene (the trp aporepressor) cannot bind to the operator, in dissimilarity to the lac repressor. Nevertheless, if tryptophan synthesis is nowadays, the aporepressor and the tryptophan molecule join together to form an active repressor complex that binds to the operator. When the external supply of tryptophan is depleted (or reduced substantially), the operator becomes exposed, and transcription begins. This type of on-off mechanism – activation of an aporepressor past the product of the biosynthetic pathway – has been observed in other biosynthetic pathways.

When the trp operon is de-repressed, which is commonly the case unless the concentration of tryptophan in the medium is very high, the optimal concentration of tryptophan is maintained by a modulating system in which the enzyme concentration varies with the concentration of tryptophan. This modulation is affected past:

1.: Premature termination of transcription before the first structural gene is reached; and
2.: Regulation of the frequency of premature termination by the concentration of tryptophan.

Located betwixt the v′ cease of the trp mRNA molecule and the start codon of the trpE cistron is a 162-base of operations segment chosen the leader (a general term for such regions). Within the leader is a sequence of bases (bases 123 through 150) with regulatory activeness. Afterward initiation of mRNA synthesis, virtually mRNA molecules are terminated in this region (except in the complete absenteeism of tryptophan), yielding a short RNA molecule consisting of only 40 nucleotides and terminating before the structural genes of the operon. This region in which termination occurs is a regulatory region called the attenuator. The base sequence around which termination occurs (Figure 24-4) has the usual features of a transcription termination site – namely, a possible stem-and-loop configuration in the mRNA followed by a sequence of viii AU pairs.

The leader sequence has an AUG codon that is in-stage with a UGA stop codon; together these start-stop signals encode a polypeptide of 14 amino acids. The leader sequence has an interesting characteristic – at positions 10 and 11 are ii side by side tryptophan codons.

Premature termination of mRNA synthesis is mediated through translation of the leader peptide. The two tryptophan codons make translation of the leader polypeptide sequence quite sensitive to the concentration of charged tRNA^Trp. If tryptophan is limiting, there will be insufficient charged tRNA^Trp and translation will suspension at the tryptophan codons. Thus biosynthesis of trp depends on ii characteristics of cistron regulation in bacteria:

1.: Transcription and translation are coupled;
two.: Base-pair formation in mRNA is eliminated in any segment of the mRNA that is in contact with the ribosome.

Effigy 24-five shows that the finish of the trp leader peptide is in segment one and a ribosome is in contact with almost 10 bases in the mRNA past the codons being translated. When the last codons are beingness translated, segments 1 and 2 are not paired. In a coupled transcription–translation organization, the leading ribosome is non far behind the RNA polymerase molecule. Thus, if the ribosome is in contact with segment 2, when synthesis of segment 4 is beingness completed, then segments iii and 4 are complimentary to form the duplex region three–iv without segment 2 competing for segment three. The presence of the 3–4 stem-and-loop configuration allows termination to occur when the terminating sequence of seven U's is reached.

If exogenous tryptophan is not present or is present in very small amounts, the concentration of charged tRNA^Trp will be limiting, and occasionally a translating ribosome will exist stalled for an instant at the tryptophan codons. These codons are located sixteen bases before the beginning of segment two. Thus, segment 2 will be free before segment 4 has been translated and the 2–iii duplex volition form. In the absenteeism of the 3–4 stalk-and-loop, termination will not occur and the complete mRNA molecule volition be made, including the coding sequences for the trp genes. Thus once once again, if tryptophan is present in excess, termination occurs and lilliputian enzyme is synthesized; if tryptophan is absent, at that place is no termination and the enzymes are made. At intermediate concentrations, the frequency of ribosome pausing will be such equally to maintain the optimal concentrations of enzymes. This tryptophan regulatory machinery is called attenuation and has been observed for several amino acid biosynthetic operons, e.g., histidine and phenylalanine. Some bacterial operons are regulated solely past attenuation without repressor–operator interactions.

Temporal mRNA Regulation in Phage Systems (meet website).

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trp Operon and Attenuation

Paul Gollnick , in Encyclopedia of Biological Chemistry, 2004

The E. coli trp Operon: Attenuation Based on Translation of a Leader Peptide

The E. coli trpEDCBA operon encodes the enzymes required to synthesize L-tryptophan from chorismic acid. Transcription of the trp operon is regulated in response to changes in intracellular tryptophan levels. When the cells contain adequate amounts of tryptophan, for example when it is nowadays in the growth medium, transcription of the operon is downwardly-regulated. In contrast, when tryptophan is limiting, the trp operon is actively transcribed in order to express the enzymes required for its synthesis. Initiation of transcription is regulated by the trp repressor, a Deoxyribonucleic acid-binding protein encoded by the trpR gene. In add-on, after transcription has initiated, the elongating transcription complex is subject to regulation by attenuation. Together, repression (80-fold) and attenuation (viii-fold) serve to allow ∼600-fold overall control of transcription of the trp operon in response to diverse levels of tryptophan availability.

Attenuation Control of the E. coli trp Operon

The E. coli trp operon contains a 162 bp leader region prior to the start of the trpE coding sequence. The trp leader transcript contains several inverted repeats, composed of the segments labeled 1–four in Figure 1, that tin form 3 different overlapping base-paired RNA secondary structures. These structures include an intrinsic transcription terminator (three:iv), an overlapping antiterminator (2:three), and a pause construction (1:2). In add-on, the leader transcription contains a minor open reading frame (ORF) that encodes a 14-amino acid leader peptide, which contains ii critical tandem UGG Trp codons. The cell's ability to efficiently interpret these two Trp codons determines which RNA structure forms in the nascent leader transcript, which in turn controls whether transcription halts in the leader region or continues into the construction genes of the operon.

Shortly subsequently transcription initiates from the trp promoter, the 1:2 pause hairpin forms (Figure one). This structure signals RNA polymerase to suspension transcription after nucleotide 92. This pausing of RNA polymerase is a critical feature of the attenuation machinery considering it allows time for a ribosome to initiate translation of the leader peptide. When the ribosome begins translating the leader peptide this releases the paused RNA polymerase to resume transcription. Transcription and translation are now coupled, with the ribosome closely following RNA polymerase. This state of affairs is essential to allow events involving the ribosome to affect transcription past the associated RNA polymerase.

At this point in that location are two possible pathways for the attenuation mechanism to follow depending on the level of tryptophan in the cell. The pick depends on how efficiently the tandem Trp codons in the leader peptide are translated. This efficiency reflects the availability of aminoacylated tRNA^Trp in the cell. Under atmospheric condition where tryptophan is limiting, the amount of charged tRNA^Trp is low. As a result of this low concentration of tryptophanyl-tRNA^Trp translation of the tandem Trp codons is inefficient and the ribosome stalls at 1 of these two codons. The associated RNA polymerase continues to transcribe through the trp leader region and transcription and translation become uncoupled. Equally RNA polymerase proceeds through segments ii and 3 of the leader region, the antiterminator structure (2:three) forms, which prevents formation of the overlapping intrinsic terminator (three:4) structure. Hence transcription continues through the leader region and into the trp structural genes.

When tryptophan is plentiful, the level of charged tRNA^Trp in the cell is high. This allows efficient translation of the tandem Trp codons and hence the ribosome proceeds rapidly to the end of the leader peptide. When the ribosome reaches the leader peptide cease codon, it covers part of RNA segment 2 and thus prevents formation of the two:3 antiterminator structure as transcription gain. This frees RNA segment three to base of operations pair with segment 4 and form the terminator. Under these conditions transcription terminates in the leader region prior to the trp structural genes, which are therefore non expressed.

In this attenuation mechanism the regulatory betoken is the level of charged tRNA^Trp and the sensory event is the efficiency with which the ribosome can translate the tandem Trp codons in the leader peptide. This arrangement tin easily be adapted to regulate other bacterial amino acid biosynthetic operons. The only needed modification is to change the identity of the critical codons in the leader peptide, which controls the amino acid the organization will respond to. There are numerous examples of such adaptations of this attenuation mechanism, especially in enteric bacteria, including the his, phe, and leu operons, which contain seven His, seven Phe and four Leu codons in their respective leader peptides.

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Gene Expression in Bacterial Systems: The trp Operon and Attenuation

P. Gollnick , in Encyclopedia of Biological Chemistry (2nd Edition), 2013

Attenuation Control of the E. coli trp Operon

The East. coli trp operon contains a 162-nucleotide leader region prior to the start of the trpE coding sequence. There are several inverted repeats in the trp leader mRNA transcript, which are equanimous of the segments labeled 1–iv in Effigy 1 . These segments can base-pair to form 3 different overlapping base-paired RNA secondary structures ( Figure one ). These structures include an intrinsic transcription terminator (3:4), an overlapping antiterminator (two:3), and a pause construction (1:ii). In addition, the leader transcription contains a pocket-size open reading frame (ORF), which encodes a xiv-amino-acid leader peptide. There are two critical tandem UGG Trp codons within this ORF. How efficiently these two Trp codons can be translated determines which RNA structure forms in the nascent leader transcript, which in plow controls whether transcription halts in the leader region or continues into the construction genes of the operon.

Soon afterwards transcription initiates from the trp promoter, the 1:ii pause hairpin forms in the nascent RNA ( Figure 1 ). This construction signals RNAP to suspend transcription after nucleotide 92. Pausing of RNAP is a critical feature of the attenuation mechanism because information technology allows time for a ribosome to initiate translation of the leader peptide. When a ribosome begins translating the leader peptide, this releases the paused RNAP to resume transcription. Transcription and translation are now coupled, with the ribosome closely post-obit RNAP. This situation is essential to allow events involving the ribosome to bear on transcription by the associated RNAP.

At this point, there are two possible pathways for the attenuation mechanism to follow depending on the level of tryptophan in the cell. The choice depends on how efficiently the tandem Trp codons in the leader peptide are translated. This efficiency reflects the availability of aminoacylated tRNA^Trp in the cell. Under conditions where tryptophan is limiting, the corporeality of charged tRNA^Trp is depression. Equally a result of the low concentration of tryptophanyl-tRNA^trp, translation of the tandem Trp codons is inefficient and the ribosome stalls at one of these two codons. The associated RNAP continues to transcribe through the trp leader region and transcription and translation become uncoupled. As RNAP continues transcribing through segments two and 3 of the leader region, the antiterminator construction (ii:3) forms, which prevents formation of the overlapping intrinsic terminator (iii:iv) structure. Hence, transcription continues through the leader region and into the structural genes of the operon which encode the tryptophan biosynthetic enzymes.

When tryptophan is plentiful, the level of charged tRNA^Trp in the cell is high. This situation allows efficient translation of the tandem Trp codons and hence the ribosome gain rapidly to the cease of the leader peptide. When the ribosome reaches the leader peptide stop codon, it covers office of RNA segment two and prevents formation of the 2:iii antiterminator structure as transcription proceeds. Thus, RNA segment 3 is free to base-pair with segment 4 and form the transcription terminator. Under these weather condition, transcription terminates in the leader region prior to the trp structural genes, which are therefore non expressed.

In this attenuation mechanism the regulatory bespeak is the level of aminoacylated tRNA^Trp and the sensory outcome is the efficiency with which the ribosome tin can translate the tandem UGG Trp codons in the leader peptide. This arrangement tin easily be adapted to regulate other amino acid biosynthetic operons. The just modification that is needed is to change the identity of the critical codons in the leader peptide, which controls the response of the system to a specific amino acrid. There are numerous examples of such adaptations of this attenuation mechanism, particularly in enteric leaner, including the his, phe, and leu operons, which contain 7 His, seven Phe, and four Leu codons in their corresponding leader peptides.

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Transcription | Expression of the Bacterial L-Trp Regulon☆

Luis R. Cruz-Vera , in Encyclopedia of Biological Chemistry (Third Edition), 2021

Sensing L-Trp Related Metabolites

Pseudomonas aeruginosa has its trp operon genes dispersed in 3 main genetic subunits. P. aeruginosa do not express the TrpR protein. The majority of the trp operon genes are under the control of the trpL regulatory gene, which senses the accumulation of uncharged tRNA^Trp (Fig. 4(A)). Withal, P. aeruginosa expresses a regulatory protein named TrpI, which instead of detecting gratuitous L-Trp and blocking the expression of genes, this regulatory protein detects indoleglycerol phosphate (InGP), inducing the expression of the trpA and trpB genes that constitute the Tryptophan synthase complex (Fig. 4(B)) (Manch and Crawford, 1982). InGP is an intermediate metabolite of the 50-Trp synthesis pathway, which Tryptophan synthase A utilizes to catalyse the germination of indole (Fig. 1). When L-Trp concentrations are low, the aggregating of uncharged tRNA^Trp induces the expression of the first four enzymes of the 50-Trp anabolic pathway, which results in the accumulation of the InGP intermediate (Fig. 4(A) , <10 µM). InGP interacts with TrpI monomers inducing dimer formation. The dimer binds a promoter region located between the trpI gene and the trpB-A operon, which are transcribed in reverse directions (Fig. 4(B) , >x µM InGP) (Chang and Crawford, 1990). The interaction of the TrpI-InGP circuitous with its promoter region induces the transcription of the trpB-A operon and represses the transcription of the trpI gene (Fig. 4(B) , >x µM InGP) (Chang and Crawford, 1990). At sufficient or excessive L-Trp concentrations, the bulk of the trp operon genes are repressed because of the low concentration of uncharged tRNA^Trp (Fig. 4(A), ≈/>ten µM). Under these conditions the concentration of InGP is reduced, the absence of InGP then reduces the interaction of the TrpI regulator with the promoter region of the trpI gene and trpB-A operon (Fig. 4(B), ≈/<10 µM InGP). These terminal events decrease the transcription of the trpB-A operon and increment the transcription of the trpI gene (Fig. 4(A), ≈/<10 µM InGP). Therefore, in Pseudomonas the regulation of the trp operon genes is decoupled, for which aggregating of InGP, perhaps produced from other sources, could be utilized to produce L-Trp without the demand to synthesize proteins required for the initial steps in the synthesis of Fifty-Trp.

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Attenuation, Transcriptional

T.1000. Henkin , in Encyclopedia of Genetics, 2001

Terminator Proteins

Expression of the Bacillus subtilis trp operon is controlled past TRAP, an unusual RNA binding poly peptide. In the presence of tryptophan, TRAP binds to the trp leader RNA and prevents formation of an antiterminator structure, thereby permitting formation of the competing intrinsic terminator. TRAP assembles into an 11-subunit symmetrical band, with 11 molecules of tryptophan spaced between the TRAP monomers. The RNA appears to wrap around the outside of the TRAP ring, with contacts betwixt each monomer and GAG/UAG repeats in the RNA binding site. TRAP oligomerization is tryptophan-independent, but RNA bounden requires tryptophan, suggesting that tryptophan controls TRAP activity by causing a conformational change that is required for binding to its RNA target site. The B. subtilis pyr arrangement is also regulated by binding of a regulatory poly peptide to the RNA leader region to mediate transcription termination. In this instance, the regulator, PyrR, binds in the presence of UMP, an end production of pyr operon expression. The target site for PyrR is a complex construction. Binding to this element precludes formation of an antiterminator structure, which competes with the attenuator. Thus, PyrR causes termination by stabilization of an anti-antiterminator. The trp and pyr systems are similar in that the default land is readthrough of the attenuator in the absence of the end-production of expression of the operon, then that transcription will be prevented only if the required metabolite is present.

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Tryptophan Operon

C. Yanofsky , in Encyclopedia of Genetics, 2001

Regulation of Expression of the trp Operon of Escherichia coli

The five structural genes of the trp operon are preceded past a transcription regulatory region consisting of a promoter/operator, at which transcription initation is regulated, and a transcribed leader segment, within which transcription termination is regulated. Initation at the trp promoter is regulated by the tryptophan-activated trp repressor protein; the extent of repression varies in response to changes in the intracellular concentration of costless tryptophan. Repression regulates operon expression over near an 80-fold range. Polymerase molecules that have initiated transcription at the trp promoter and escaped repression are subject to a second regulatory machinery, transcription attenuation. The latter mechanism determines whether or non transcription volition terminate at a site located in the distal portion of the leader region. This decision is influenced by the intracellular concentration of tryptophan-charged tRNA^Trp. When the Trp-tRNA^Trp concentration is high, transcription terminates in the leader region. When tRNA^Trp is mostly uncharged, which occurs when cells experience a severe tryptophan deficiency, termination is avoided and transcription proceeds to the terminate of the operon. Transcription attenuation in the trp operon of E. coli regulates transcription of the structural genes of the operon over about an eightfold range. The combined action of repression and attenuation regulates transcription of the structural genes of the operon over about a 600-fold range. There is an internal promoter located in the distal portion of trpD (Figure 1). Transcription initiation at this promoter is unregulated and proceeds at a frequency less than 10% that owing to the principal promoter. Tandem sites of transcription termination are located post-obit the trpA structural gene; the start is poly peptide-factor-independent, a and then-called intrinsic terminator, while the second required the protein Rho. Completion of transcription of the operon yields a polycistronic messenger RNA. Ribosomes tin can initiate translation at any of the five major ribosome binding sites on this polycistronic messenger.

The trp promoter region of E. coli contains 3 operators that tin can bind trp repressor. Operator-spring repressor inhibits transcription initiation. The trp repressor also regulates transcription initiation in several other operons concerned with tryptophan metabolism. The three-dimensional structures of the trp aporepressor (aporepressor lacks spring tryptophan), the trp repressor, and the trp repressor–operator complex, have been adamant. These structures have revealed the features of this protein that are responsible for its activation by tryptophan and its recognition of specific operators.

The transcribed leader region of the trp operon of East. coli is about 160 bp in length. As mentioned, this genetic segment encodes an mRNA segment that can cause transcription termination in the leader region. The transcript of the leader region can fold to form iii RNA structures, termed terminator, antiterminator, and transcription pause structure. The terminator and antiterminator are culling RNA structures, i.e., they take a sequence of nucleotides in common, thus either, just not both, can exist at once. When cells are scarce in charged tRNA^Trp the antiterminator forms; this precludes formation of the terminator. When cells take adequate levels of charged tRNA^Trp, the terminator forms and transcription terminates in the leader region. A deficiency of charged tRNA^Trp is sensed during attempted translation of tandem Trp codons in a 14-residue leader peptide coding region, trpL, located near the five′ terminate of the trp operon transcript. Coupling of transcription and translation, essential to this mechanism of attenuation, is achieved past the formation of the transcription pause structure, located near the 5′ end of the transcript. Polymerase pausing allows a ribosome to bind to the transcript and initiate synthesis of the leader peptide. The move of this ribosome and so releases the paused transcription complex, and transcription and translation proceed in unison.

Two of the trp polypeptides, the products of genes trpE and trpA, lack tryptophan, therefore they are synthesized preferentially during astringent tryptophan starvation. An additional regulatory characteristic, translational coupling, insures equimolar synthesis of the polypeptide products of two pairs of adjacent genes, trpE and trpD, and trpB and trpA. As mentioned, the products of these genes form enzyme complexes. The enzyme complex catalyzing the first two reactions in the pathway is feedback-inhibited past tryptophan. The tryptophan bounden site is located in the TrpE polypeptide.

The apply of ii transcription regulatory mechanisms and feedback inhibition of anthranilate synthase activity allows E. coli to regulate tryptophan biosynthesis efficiently in response to changes in the availability of tryptophan and the rate of protein synthesis.

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The Folding of Proteins and Nucleic Acids

50. Liu , A.1000. Gronenborn , in Comprehensive Biophysics, 2012

3.eight.3.11 TrpR

The trp repressor (trpR) binds the operator region of the trp operon and prevents the initiation of transcription. Its smallest functional unit of measurement is a dimer, ¹⁵⁰ although tetrameric and higher order species are also observed. ^151,152 Under extreme atmospheric condition (30% isopropanol), an infinite crystalline 3-D supramolecular array was constitute, ²⁴ peradventure involving domain swapping.

Whether or not the trpR dimer constitutes a true domain-swapped example is unclear considering a highly intertwined structure was observed and no monomeric homolog is available. Each single polypeptide chain in trpR dimer is composed of six α helices (A–F) in a relative closed conformation (Figure fourteen(a)). ²³ In the crystalwide assembly, helices C–E rearrange and grade a very long, single helix, resulting in polypeptide termini that are separated past a large distance (Effigy 14(b)). 2 such dimers come together and create a substructure (half of the tetramer) very similar to that seen in the dimer (Effigy 14(c)). Therefore, the dimer-like structure is formed past segments of four different polypeptide chains, two providing intertwined Due north-last regions and two providing C-terminal regions, ²⁴ with the polymer constituting a branched aggregate rather than a daisy chain-blazon linear i. Given that the polymeric grade was crystallized in the presence of a significant amount of booze, its relevance to whatever physiological state is unclear. Indeed, it is well-known that alcohols increase the helical content of flexible peptides and/or destabilize tertiary structures significantly. ^24,153,154

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Chlamydia trachomatis

Huizhou Fan , Guangming Zhong , in Molecular Medical Microbiology (Second Edition), 2015

Variation in the Tryptophan Synthase

Human cells are auxotrophic to tryptophan. In contrast, many microbes are capable of synthesizing tryptophan de novo. Enzymes required for synthesizing tryptophan are encoded on the trp operon. The trp operon in C. trachomatis contains three open up reading frames (ORFs), designated trpR, trpB and trpA, which encode proteins TrpR, TrpB and TrpA, respectively. Bacterial tryptophan synthase is generally comprised of two TrpA polypeptides (α_ii) and two TrpB polypeptides (β₂) in an α₂β₂ configuration. The synthase is a bifunctional enzyme, in which the α subunits convert indole glycerol 3-phsophate (IGP) into indole and glyceraldehyde-3-phosphate, and the β subunits add together a serine residue to indole, thereby yielding tryptophan. The two sequential reactions occur in a highly cooperative manner [70–72]; accordingly, removal of one pair of subunits is sufficient to crusade a near complete halt in the enzyme activity of the other pair of subunits [73].

Genetic and biochemical analyses have shown substantial changes in the C. trachomatis tryptophan synthase, as compared with the enzyme in other leaner. C. trachomatis TrpB shares a relatively high sequence identity with E. coli TrpB (54%), including all disquisitional residues at the active site of the enzyme. In contrast, the caste of amino acid conservation between C. trachomatis TrpA and E. coli TrpA is very depression (27%) [74]. Furthermore, in all the genital serovars, in-frame deletions have resulted in the deletion of a number of critical residues in TrpA, leading to loss of the ability to catalyse the formation of indole from IGP; however, the highly mutated TrpA is still required for the generation of tryptophan by TrpB. In ocular C. trachomatis serovars, there are additional mutations causing frame-shift, leading to complete inactivation of tryptophan synthesis activeness; a complete loss of trpBA has too been found in an ocular strain. Thus, there is a complete correlation between tryptophan synthesis and C. trachomatis tissue tropism: while all genital serovars can synthesize tryptophan using indole as a substrate, ocular serovars cannot.

In addition to TrpA and TrpB ORFs, bacterial trp operons also deport ORFs for enzymes catalysing a series of iv additional reactions starting from chorismate to IGP [75,76]. None of these boosted genes is plant in the C. trachomatis genome. Thus, C. trachomatis can satisfy the requirement for tryptophan by either save from the host cell or synthesizing this amino acid from indole as the merely starting fabric. Since human cells cannot produce indole, the source of tryptophan precursor is presumably other microbes, which are in loftier abundance in the genital tract, but non in the eye. This may explicate why genital C. trachomatis serovars have retained a functional tryptophan synthase and the ocular serovars have not.

The tryptophan acquisition mechanism in C. trachomatis is linked to the pathogen'due south power to tolerate the antichlamydial cytokine interferon-γ. Human interferon-γ causes tryptophan starvation by inducing the expression of indoleamine-ii,3-dioxygenase (IDO), which catalyses the degradation of tryptophan. While indole as well as tryptophan can reverse the inhibitory effect of human interferon-γ on C. trachomatis genital serovars, only tryptophan but not indole is able to alleviate the growth inhibition in the ocular strains.

Paradoxically, loss of the tryptophan biosynthesis pathway actually contributes to the pathogenesis of ocular strains. Tryptophan starvation can cause chlamydiae to enter persistence, which is well documented in cell civilization systems [77–79]. Chlamydial persistence is idea to exist responsible for trachoma and other chronic middle manifestations. Without the power to synthesize tryptophan, ocular strains are likely to suffer more than astringent tryptophan impecuniousness in the presence of man interferon-γ, and are more forcefully driven to enter persistence, as compared to genital strains.

TrpR is a transcription regulator of the trp operon. The function of TrpR is regulated by the bioavailability of gratuitous tryptophan in the microbial jail cell [76,lxxx–83]. When bacteria are grown in media rich in tryptophan, the homodimeric TrpR is bound to two molecules of tryptophan, forming an active autorepressor complex, which binds to the trp operator, a DNA sequence near a promoter element recognized by the σ factor of the RNA polymerase. Occupation of the trp operator by the autorepressor prevents the RNA polymerase from binding to the trp promoter and initiating trp RNA synthesis. When the concentration of tryptophan drops, tryptophan dissociates from the autorepressor, which in turn dissociates from the trp operator, allowing RNA polymerase to synthesize the trp mRNA [84]. It has been demonstrated that TrpR from C. trachomatis serovar D and L2 binds to the trp promoter in vitro [85,86], and furthermore, TrpR represses transcription of the trp operon in a tryptophan-dependent style in vitro [85]. Every bit concluding proof that TrpR acts equally a negative regulator of transcription in C. trachomatis, the transcription is no longer regulated by tryptophan in a C. trachomatis L2 variant containing a frameshift mutation in trpR [86].

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