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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The cysK gene of Escherichia coli K-12 encoding O-acetylserine sulphydrylase A, was cloned and its nucleotide sequence, together with that of the flanking regions, was determined. The deduced amino acid sequence of the carboxy-terminal moiety of O-acetylserine sulphydrylase A shows significant similarity to the amino acid sequence of tryptophan synthase beta chain from several organisms. This sequence similarity is likely to reflect the structural homologies of substrates shared by both enzymes. This may indicate that these proteins, although catalysing different reactions in different metabolic pathways, have evolved from a common ancestral gene.
Mol Microbiol 1988 Nov
PMID:Phylogeny of metabolic pathways: O-acetylserine sulphydrylase A is homologous to the tryptophan synthase beta subunit. 306 11

The G + C content of DNA varies widely in different organisms, especially microorganisms. This variation is accompanied by changes in the nucleotide composition of silent positions in codons. (Silent positions are defined and explained in the text). These changes are mostly neutral or near neutral, and appear to result from mutation pressure in the direction of increasing either A + T (AT pressure) or G + C (GC pressure) content. Variations in G + C content are also accompanied by substitutions at replacement positions in codons. These substitutions produce changes in the amino acid content of homologous proteins. The examples studied were genes for 13 mitochondrial proteins in five species, and A and B genes for bacterial tryptophan synthase in four species. In microorganisms, varying AT and GC mutational pressures, presumably resulting from shifts in the DNA polymerase system, exert strong effects on molecular evolution by changing the G + C content of DNA. These effects may be greater than those of random drift. The effects of GC pressure on silent substitutions in the systems examined are several times as great as the effects on replacement substitutions. GC pressure is exerted on noncoding as well as coding regions in mitochondrial DNA. This is shown by the close correlation (correlation coefficient, 0.99) of the G + C content of the noncoding D loop of mitochondria with the G + C content of silent positions in the corresponding mitochondrial genes.
J Mol Evol 1986
PMID:Silent nucleotide substitutions and G + C content of some mitochondrial and bacterial genes. 310 17

We have determined the DNA sequence of the two adjacent genes for the alpha and beta chains of tryptophan synthase in Pseudomonas aeruginosa, along with 34 5'-flanking and 799 3'-flanking base pairs. The gene order is trpBA as predicted from earlier genetic studies, and the two cistrons overlap by 4 bp; a ribosome binding site for the second gene is evident in the coding sequence of the first gene. We have also determined the location of three large deletions eliminating portions of each gene. A detailed comparison of the deduced P. aeruginosa amino acid sequence with those published for E. coli, Bacillus subtilis, and Saccharomyces cerevisiae shows much similarity throughout the beta and most of the alpha subunit. Most of the residues implicated by chemical modification or mutation as being critical for enzymatic activity are conserved, along with many others, suggesting that three-dimensional structure has remained largely constant during evolution. We also report the construction of a recombinant plasmid that overproduces a slightly modified alpha subunit from P. aeruginosa that can form a functionally effective multimer with normal E. coli beta 2 subunit in vivo.
Mol Biol Evol 1986 May
PMID:Nucleotide sequence of the genes for tryptophan synthase in Pseudomonas aeruginosa. 312 51

A cosmid bank of Methanococcus voltae DNA was obtained in Escherichia coli after ligation of partially HindIII-digested M. voltae DNA in the HindIII site of the transferable cosmid pVK100. The bank was used to perform complementation experiments with E. coli auxotrophic mutants. Five cosmids complementing trpA shared three adjacent HindIII fragments of 2.1, 2.3 and 14 kb. Two of these cosmids also complemented trpD and carried an additional 4.2 kb HindIII fragment. The trpA- and trpD- complementing regions were more precisely localized using Tn5 mutagenesis. A 1.7 kb PstI fragment, cloned into pUC9 in both orientations, was responsible for the trpA complementation. This fragment was sequenced and an open reading frame (ORF) of 852 nucleotides (ORFtrpA) encoding a 284 amino acid polypeptide of mol. wt. 31,938 was found. The amino acid sequence was compared with that of the alpha subunit of tryptophan synthase (trpA gene product) from nine eubacterial species and to the N-terminal part of the tryptophan synthase of Saccharomyces cerevisiae (TRP5 gene product). Similarity varied from 24% (Brevibacterium lactofermentum) to 35% (S. cerevisiae). The nucleotide sequence of the region upstream from M. voltae ORFtrpA was determined and revealed the presence of an ORF of 1227 nucleotides (ORFtrpB) encoding a 409 amino acid polypeptide of mol. wt. 44,634. The polypeptide sequence was similar to the beta subunit of tryptophan synthase (trpB gene product) from six eubacterial species and to the C-terminal part of the tryptophan synthase of S. cerevisiae. Similarity varied from 49% (S. cerevisiae, B. lactofermentum) to 58% (Pseudomonas aeruginosa). This high conservation supports the hypothesis of a common ancestor for the trpA and trpB genes of archaebacteria, eubacteria and eucaryotes. M. voltae ORFtrpA and ORFtrpB, which are transcribed in the same direction, are separated by a 37 bp AT-rich region. Immediately upstream from ORFtrpB, the 3' end of an ORF homologous to E. coli and Bacillus subtilis trpF was found. As the trpD-complementing region was located upstream from the trpFBA sequenced region, the organization of trp genes in the archaebacterium might thus be trpDFBA. Such an organization resembles that of enteric eubacteria, in which the trpEDCFBA genes are grouped in a single operon. However, M. voltae ORFtrpA and ORFtrpB do not overlap, in contrast with what is found in most eubacteria.
Mol Gen Genet 1988 Nov
PMID:Cloning of the trp genes from the archaebacterium Methanococcus voltae: nucleotide sequence of the trpBA genes. 314 17

The equilibrium and kinetic properties for the urea-induced unfolding of the alpha subunit of tryptophan synthase from Escherichia coli, Salmonella typhimurium, and five interspecies hybrids were compared to determine the role of protein folding in evolution. The parent proteins differ at 40 positions in the sequence of 268 amino acids, and the hybrids differ by up to 15 amino acids from the Escherichia coli alpha subunit. The results show that all the proteins follow the same folding mechanism and are consistent with a previously proposed hypothesis [Hollecker, M., & Creighton, T. E. (1983) J. Mol. Biol. 168, 409; Krebs, H., Schmid, F. X., & Jaenicke, R. (1983) J. Mol. Biol. 169, 619] that the folding mechanisms are conserved in homologous proteins. Analysis of the kinetic data suggests that the 15 positions at which the parent proteins differ in the amino folding unit, residues 1-188, do not play a role in a rate-limiting step in folding that has been previously identified as the association of the amino and carboxyl folding units [Beasty, A. M., Hurle, M. R., Manz, J. T., Stackhouse, T. S., Onuffer, J. J., & Matthews, C. R. (1986) Biochemistry 25, 2965]. One or more of the 25 positions at which the parent proteins differ in the carboxyl folding unit, residues 189-268, do appear to play a role in this same rate-limiting step.
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PMID:Folding of homologous proteins: conservation of the folding mechanism of the alpha subunit of tryptophan synthase from Escherichia coli, Salmonella typhimurium, and five interspecies hybrids. 328 27

Included in a series of monoclonal antibodies obtained after immunization with the native holo beta 2 subunit of tryptophan synthase of Escherichia coli (EC 4.2.1.20), are some that interact preferentially with a denatured state of the antigen (Friguet et al., 1984). A study of the equilibrium and kinetic characteristics of the interaction of one of these antibodies with native apo beta 2 (i.e. free of pyridoxal 5'-phosphate) and with one of its proteolytic domains is reported here. The antibody is shown to interact strongly with the isolated domain in accordance with a simple equilibrium. In the presence of native beta 2, the antibody binds exclusively to the dissociated beta-monomer. The interaction of this antibody with native apo beta 2 is used to determine the equilibrium and kinetic constants of the monomer-dimer equilibrium. The values obtained are 4.5 X 10(-8) M for the equilibrium constant and 7.9 X 10(-3) s-1 for the rate constant of the dissociation of apo beta 2 into beta-monomers.
J Mol Biol 1987 Sep 05
PMID:Kinetics of the spontaneous transient unfolding of a native protein studied with monoclonal antibodies. Monomer/dimer transition in the tryptophan-synthase beta 2 subunit. 331 67

During its folding, the polypeptide chain of the beta 2 subunit of Escherichia coli tryptophan synthase (L-serine hydrolyase (adding indole) EC 4.2.1.20) undergoes dimerization. To decide whether this dimerization precedes or follows the formation of the native, functional, tertiary structure of the polypeptide chain, the kinetics of renaturation of beta 2 are studied by monitoring both the regain of native conformation and the dimerization. Dimer formation is followed by the increase of the fluorescence polarization, or by energy transfer between a fluorescence donor and a fluorescence acceptor, which occur upon association of adequately labelled beta chains. Renaturation is followed by the regain of functional properties of beta 2, i.e. its ability to bind pyridoxal-5'-phosphate or to form a fluorescent ternary complex with this coenzyme and L-serine. It is shown that for beta 2 the dimerization obeys first-order kinetics, presumably because it occurs rapidly after a rate-limiting isomerization of the monomer. The dimerization is followed by another isomerization, taking place within the dimer, which leads to the formation of the pyridoxal-5'-phosphate binding site. Still another, slow, isomerization reaction involving the F1 (N-terminal) domain completes the renaturation. With a modified form of beta 2 (trypsin-nicked beta 2) where this isomerization of F1 can be made to occur before the dimerization, the dimer is also shown to appear before the functional properties. It is concluded that a non-native dimer indeed exists as an obligatory intermediate on the folding pathway of nicked beta 2 and of beta 2, and that interdomain interactions are needed to force the polypeptide chains into their native conformations.
J Mol Biol 1985 Apr 20
PMID:Kinetics and importance of the dimerization step in the folding pathway of the beta 2 subunit of Escherichia coli tryptophan synthase. 389 16

Recent studies with monoclonal antibodies directed against different epitopes of the beta 2-subunit of Escherichia coli tryptophan synthase have shown that some of these antibodies bind rapidly in solution to the native protein; others bind very slowly to native beta 2 in solution while they recognize quite rapidly this antigen adsorbed on a microtitration plate. In the present work, an enzyme-linked immunosorbent assay competition test with either the native or a denatured form of the antigen has been developed. It allowed us to show that the rapidly binding antibodies recognize epitopes present on the native protein while those which react very slowly in solution bind preferentially to the denatured form of the protein. These results prompted us to emphasize how important it is, in the characterization of antibodies, to ascertain that the antigen they recognize remains native in the specificity test.
Mol Immunol 1984 Jul
PMID:Some monoclonal antibodies raised with a native protein bind preferentially to the denatured antigen. 620 56

The interaction of Trp repressor protein with partial trp operators was studied in vitro and in vivo. At high ratios of protein to DNA, Trp holorepressor formed stable complexes with DNA molecules containing half operators. When plasmids conferring the capacity to hyperproduce Trp repressor were present in trpOc strains of Escherichia coli, repression of downstream tryptophan synthase occurred. Palindromicity of the trp operator may facilitate stable interaction with Trp repressor, but this attribute need not be regarded as a critically essential structural feature. Sufficient information for the recognition by Trp repressor protein of an appropriate target resides within a DNA sequence of approximately ten base-pairs.
J Mol Biol 1983 Nov 15
PMID:Studies on the interaction of Trp holorepressor with several operators. Evidence that the target need not be palindromic. 635 17

All five tryptophan biosynthetic genes of Saccharomyces cerevisiae were unified on plasmid pME554, which is based on 2 micrometer DNA and pBR322 sequences allowing for autonomous replication in yeast and E. coli. Homologous and heterologous expression of this artificial yeast TRP-gene cluster was studied. Plasmid pME554 allowed for nearly normal growth of a yeast strain bearing auxotrophic mutations in all five TRP-genes. The plasmid-borne genes TRP2 to TRP5 were expressed and regulated normally in the frame of the general control. Gene TRP1, carried on an EcoRI/Bg/II fragment lacking the ARS1 function, was expressed poorly and did not respond to the general control like the chromosomally-borne TRP1 gene. Plasmid pME554 allowed for poor growth of E. coli strain W3110 tna- delta trpEA2 on minimal medium. Marked stimulation was observed, however, when anthranilic acid or indole were added. Accordingly, poor expression of the first Trp-enzyme anthranilate synthase and the last enzyme tryptophan synthase was found, whereas the other three genes were moderately well expressed in E. coli.
Mol Gen Genet 1984
PMID:Expression of an artificial yeast TRP-gene cluster in yeast and Escherichia coli. 638 66


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