The subunit structure of neurospora tryptophan synthase: A reappraisal

Tsai, Hui Lien; Yang, Cheng-Fu; Tsai, Jane H. J.

doi:10.1016/s0006-291x(74)80430-4

Cited by 13 publications

(6 citation statements)

References 26 publications

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“…Instead, translation would continue into trpA (in-phase) to produce a fused trpB-trpA polypeptide. In fact, in Neurospora crassa the trpB and trpA functions do appear to reside in a single polypeptide chain (25,26). A similar fusion mechanism might also have given rise to the trpD and trpC proteins in E. coli, which each now possesses two separate enzymatic functions in single polypeptide chains.…”

Section: Cgu-ggc-gau-aaa-gac-auc-uuc-acc-guu-cac-gau-aut-uug-aaa-gca-mentioning

confidence: 99%

“…We show here that the ribosome-protected fragment spans the end of trpB as well as the beginning of trpA, and that the intercistronic region consists of only two untranslated nucleotides. J3y contrast, the known intercistronic regions in RNA bacteriophage vary in length from 25 to over 600 nucleotides; in these cases the intercistronic region may have specific functions (3).…”

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confidence: 99%

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An intercistronic region and ribosome-binding site in bacterial messenger RNA.

Platt¹,

Yanofsky²

1975

Proc. Natl. Acad. Sci. U.S.A.

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A messenger RNA fragment about 220 nucleotides long has been isolated from 2P-labeled tryptophan operon mRNA of Escherichia coli. When point mutations at the end of trpB and the beginning of trpA were introduced, the resulting nucleotide changes were found; hence the mRNA fragment must include the trpB-trpA intercistronic region. Most of the nucleotide sequences can be assigned to specilfle-lcations in the structural genes, based on the amino-acid sequences of the trpB and trpA proteins. In vitro, ribosomes bind to this piece of mRNA and protect from nuclease attack a region about 40 The nature of the "punctuation" signals governing the translation of messenger RNA is of particular importance in understanding regulation of protein synthesis. Polypeptide chain termination can be specified by any of the three "nonsense" triplet codons, UAG, UAA, or UGA (1). The principal codon used for polypeptide chain initiation is AUG (1), but it is generally believed that AUG alone is not sufficient to initiate translation. The additional signals that are required remain unknown (2). In polycistronic mRNA, "intercistronic" regions may occur between the final codon of one gene and the initiation codon of the next-several such regions from RNA bacteriophage have been sequenced (3). Studies on the histidine operon of Salnmonella typhimurium indicate that at least one nucleotide occurs between the termination codon of hiMD and the initiation codon of the following gene, hisC, but no upper limit could be placed on the size of this region (4).Recently, procedures to isolate defined segments of mRNA by specific hybridizations and refined techniques of nucleotide sequencing have made it possible to determine directly the primary structure of bacterial messenger RNA labeled in vivo with 32p (5, 6). We have used these procedures to purify and sequence a segment of tryptophan (trp) operon mRNA from Escherichia coli containing the intercistronic region between trpB and trpA (the last two genes in the operon) and overAbbreviations: Hepes(NH4), NV-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (titrated with NH40H); TKE, Tris/KCl/ 'EDTA; HG, Hepes/NH4Cl/Mg(AcO)2/dithiothreitol. 2399 lapping the structural genes on either side. The ribosomebinding approach pioneered by Steitz (7) was used to demonstrate that the "initiator" region for the trpA polypeptide is still potentially functional in this mRNA sequence. We show here that the ribosome-protected fragment spans the end of trpB as well as the beginning of trpA, and that the intercistronic region consists of only two untranslated nucleotides. J3y contrast, the known intercistronic regions in RNA bacteriophage vary in length from 25 to over 600 nucleotides; in these cases the intercistronic region may have specific functions (3). MATERIALS AND METHODSLabeling and RNA Purification. An E. coli strain merodiploid for the internal deletion trpALD102 (formerly trpAED-102) produces high levels of distal trp mRNA (8); we estimate that >100 copies of trp operon mRNA are present per cell...

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Section: Cgu-ggc-gau-aaa-gac-auc-uuc-acc-guu-cac-gau-aut-uug-aaa-gca-mentioning

confidence: 99%

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confidence: 99%

An intercistronic region and ribosome-binding site in bacterial messenger RNA.

Platt¹,

Yanofsky²

1975

Proc. Natl. Acad. Sci. U.S.A.

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“…this respect, it more closely resembles the tryptophan Tryptophan synthase (EC 4.2.1 .20, L-serine hydro-synthase of Neurospora crassa (Tsai et al 1974; lyase (adding indole)) catalyzes the final step in the Matchett and DeMoss 1975) than the enzymes of such biosynthesis of tryptophan, the conversion of indole gram-negative bacteria as Escherichia coli (Creighton 3-glycerol phosphate to tryptophan. The overall reacand Yanofsky 1966) and Pseudomonas putida (Enatsu tion can be separated into two parts: (i) the A reaction or and Crawford 197 1).…”

Section: Introductionmentioning

confidence: 97%

Studies on the association of the α and β₂ subunits of the tryptophan synthase of Bacillus subtilis

Hoch¹,

Soldau²

1981

Can. J. Microbiol.

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Studies using Sephadex gel filtration indicated that the alpha and beta 2 components of the Bacillus subtilis tryptophan synthase associate to form complexes under the appropriate conditions of buffer, pH, and temperature. Monovalent cations, glycerol, and the cofactor, pyridoxal-5'-phosphate, were required to maintain and active beta 2 component, and in turn, affected the association of the alpha and beta 2 components. Under conditions that stabilized the individual components during their purification, the affinity of the subunits for each other was weak. Under the same buffer conditions, but at the higher pH of 7.8 which the enzymatic activities are assayed, the individual components readily associated. The substrate serine appeared to affect complex formation but there was no effect from the indole moiety. When the temperature was raised from 4 to 22-25 degrees C, complex formation was observed at both pH 6.6 and 7.8. The results of these experiments are consistent with the formation of alpha beta 2 and alpha 2 beta 2 species as the associated tryptophan synthase complexes of B. subtilis.

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“…For example, the Escherichia coli tryptophan synthase is an a-P-P-a heterotetramer, whereas in filamentous fungi the genes encoding the a and / 3 subunits have fused, and the tryptophan synthase is a homodimer (Hyde et al, 1988 ;Matchett and De Moss, 1975;Tsai et al, 1974). The enzymic properties of these two tryptophan synthases are essentially identical (Tsai et al, 1974).…”

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confidence: 99%

“…The enzymic properties of these two tryptophan synthases are essentially identical (Tsai et al, 1974). Some enzymically active complexes are also complex associations of particular enzymes some which are multidomain and an example of this latter phenomenon is seen in the 2-0x0 acid dehydrogenase complex (for excellent reviews see Perham, 1991;Patel and Roche, 1990;Roche and Patel, 1989).…”

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confidence: 99%

The Molecular Biology of Multidomain Proteins Selected Examples

Hawkins

Lamb

1995

European Journal of Biochemistry

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The aim of this review is to give an overview of the contribution molecular biology can make to an understanding of the functions and interactions within multidomain proteins. The contemporary advantages ascribed to multidomain proteins include (a) the potential for metabolite channelling and the protection of unstable intermediates ; (b) the potential for interactions between domains catalysing sequential steps in a metabolic pathway, thereby giving the potential for allosteric interactions; and (c) the facility to produce enzymic activities in a fixed stoichiometric ratio. The alleged advantages in (a) and (b) however apply equally well to multi-enzyme complexes ; therefore, specific examples of these phenomena are examined in multidomain proteins to determine whether the proposed advantages are apparent. Some transcription-regulating proteins active in the control of metabolic pathways are composed of multiple domains and their control is exerted and modulated at the molecular level by protein-DNA, proteinprotein and protein -metabolite interactions. These complex recognition events place strong constraints upon the proteins involved, requiring the recognition of and interaction with different classes of cellular metabolites and macromolecules. Specific examples of transcription-regulating proteins are examined to probe how their multidomain nature facilitates a general solution to the problem of multiple recognition events. A general unifying theme that emerges from these case studies is that a basic unitary design of modules provided by enzymes is exploited to produce multidomain proteins by a complex series of gene duplication and fusion events. Successful modules provided by enzymes are co-opted to new function by selection apparently acting upon duplicated copies of the genes encoding the enzymes. In multidomain transcription-regulating proteins, former enzyme modules can be recruited as molecular sensors that facilitate presumed allosteric interactions necessary for the molecular control of transcription.

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The subunit structure of neurospora tryptophan synthase: A reappraisal

Cited by 13 publications

References 26 publications

An intercistronic region and ribosome-binding site in bacterial messenger RNA.

An intercistronic region and ribosome-binding site in bacterial messenger RNA.

Studies on the association of the α and β₂ subunits of the tryptophan synthase of Bacillus subtilis

The Molecular Biology of Multidomain Proteins Selected Examples

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The subunit structure of neurospora tryptophan synthase: A reappraisal

Cited by 13 publications

References 26 publications

An intercistronic region and ribosome-binding site in bacterial messenger RNA.

An intercistronic region and ribosome-binding site in bacterial messenger RNA.

Studies on the association of the α and β2 subunits of the tryptophan synthase of Bacillus subtilis

The Molecular Biology of Multidomain Proteins Selected Examples

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Studies on the association of the α and β₂ subunits of the tryptophan synthase of Bacillus subtilis