scholarly journals 5-Fluoroindole resistance identifies tryptophan synthase beta subunit mutants in Arabidopsis thaliana.

Genetics ◽  
1995 ◽  
Vol 140 (1) ◽  
pp. 303-313
Author(s):  
A J Barczak ◽  
J Zhao ◽  
K D Pruitt ◽  
R L Last
Keyword(s):  
Arabidopsis Thaliana ◽  
Protein A ◽  
Beta Subunit ◽  
Alpha Subunit ◽  
Anthranilate Synthase ◽  
Tryptophan Synthase ◽  
Wild Type ◽  
Recessive Mutations ◽  
The Stability

Abstract A study of the biochemical genetics of the Arabidopsis thaliana tryptophan synthase beta subunit was initiated by characterization of mutants resistant to the inhibitor 5-fluoroindole. Thirteen recessive mutations were recovered that are allelic to trp2-1, a mutation in the more highly expressed of duplicate tryptophan synthase beta subunit genes (TSB1). Ten of these mutations (trp2-2 through trp2-11) cause a tryptophan requirement (auxotrophs), whereas three (trp2-100 through trp2-102) remain tryptophan prototrophs. The mutations cause a variety of changes in tryptophan synthase beta expression. For example, two mutations (trp2-5 and trp2-8) cause dramatically reduced accumulation of TSB mRNA and immunologically detectable protein, whereas trp2-10 is associated with increased mRNA and protein. A correlation exists between the quantity of mutant beta and wild-type alpha subunit levels in the trp2 mutant plants, suggesting that the synthesis of these proteins is coordinated or that the quantity or structure of the beta subunit influences the stability of the alpha protein. The level of immunologically detectable anthranilate synthase alpha subunit protein is increased in the trp2 mutants, suggesting the possibility of regulation of anthranilate synthase levels in response to tryptophan limitation.

Characterization of tryptophan synthase alpha subunit mutants of Arabidopsis thaliana

1996 ◽  
Vol 253 (3) ◽  
pp. 353-361 ◽  
Author(s):  
E. R. Radwanski ◽  
A. J. Barczak ◽  
R. L. Last
Keyword(s):  
Arabidopsis Thaliana ◽  
Alpha Subunit ◽  
Tryptophan Synthase

scholarly journals Substitution of a Surface-Exposed Residue Involved in an Allosteric Network Enhances Tryptophan Synthase Function in Cells

2021 ◽  
Vol 8 ◽  
Author(s):  
Rebecca N. D’Amico ◽  
Yuliana K. Bosken ◽  
Kathleen F. O’Rourke ◽  
Alec M. Murray ◽  
Woudasie Admasu ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Molecular Dynamics ◽  
Active Site ◽  
Beta Subunit ◽  
Alpha Subunit ◽  
Tryptophan Synthase ◽  
Wild Type ◽  
Auxotrophic Strain ◽  
Hydrophobic Tunnel ◽  
Dynamics Simulations

Networks of noncovalent amino acid interactions propagate allosteric signals throughout proteins. Tryptophan synthase (TS) is an allosterically controlled bienzyme in which the indole product of the alpha subunit (αTS) is transferred through a 25 Å hydrophobic tunnel to the active site of the beta subunit (βTS). Previous nuclear magnetic resonance and molecular dynamics simulations identified allosteric networks in αTS important for its function. We show here that substitution of a distant, surface-exposed network residue in αTS enhances tryptophan production, not by activating αTS function, but through dynamically controlling the opening of the indole channel and stimulating βTS activity. While stimulation is modest, the substitution also enhances cell growth in a tryptophan-auxotrophic strain of Escherichia coli compared to complementation with wild-type αTS, emphasizing the biological importance of the network. Surface-exposed networks provide new opportunities in allosteric drug design and protein engineering, and hint at potential information conduits through which the functions of a metabolon or even larger proteome might be coordinated and regulated.

Molecular characterization of three mutations in katG affecting the activity of hydroperoxidase I of Escherichia coli

1990 ◽  
Vol 68 (7-8) ◽  
pp. 1037-1044 ◽  
Author(s):  
Peter C. Loewen ◽  
Jacek Switala ◽  
Mark Smolenski ◽  
Barbara L. Triggs-Raine
Keyword(s):  
Escherichia Coli ◽  
Specific Activity ◽  
Polymerase Chain Reaction Amplification ◽  
Protein A ◽  
Wild Type ◽  
Wild Type Enzyme ◽  
Gene Encoding ◽  
Reaction Amplification ◽  
Mutant Genes

Hydroperoxidase I (HPI) of Escherichia coli is a bifunctional enzyme exhibiting both catalase and peroxidase activities. Mutants lacking appreciable HPI have been generated using nitrosoguanidine and the gene encoding HPI, katG, has been cloned from three of these mutants using either classical probing methods or polymerase chain reaction amplification. The mutant genes were sequenced and the changes from wild-type sequence identified. Two mutants contained G to A changes in the coding strand, resulting in glycine to aspartate changes at residues 119 (katG15) and 314 (katG16) in the deduced amino acid sequence of the protein. A third mutant contained a C to T change resulting in a leucine to phenylalanine change at residue 139 (katG14). The Phe139-, Asp119-, and Asp314-containing mutants exhibited 13, < 1, and 18%, respectively, of the wild-type catalase specific activity and 43, 4, and 45% of the wild-type peroxidase specific activity. All mutant enzymes bound less protoheme IX than the wild-type enzyme. The sensitivities of the mutant enzymes to the inhibitors hydroxylamine, azide, and cyanide and the activators imidazole and Tris were similar to those of the wild-type enzyme. The mutant enzymes were more sensitive to high temperature and to β-mercaptoethanol than the wild-type enzyme. The pH profiles of the mutant catalases were unchanged from the wild-type enzyme.Key words: catalase, hydroperoxidase I, mutants, sequence analysis.

scholarly journals Rice OASA1D, a mutant anthranilate synthase .ALPHA. subunit gene, is an effective selectable marker for transformation of Arabidopsis thaliana

2005 ◽  
Vol 22 (4) ◽  
pp. 271-276 ◽  
Author(s):  
Makiko Kawagishi-Kobayashi ◽  
Naoto Yabe ◽  
Mizuho Tsuchiya ◽  
Sachiyo Harada ◽  
Tomoko Kobayashi ◽  
...  
Keyword(s):  
Arabidopsis Thaliana ◽  
Selectable Marker ◽  
Alpha Subunit ◽  
Anthranilate Synthase ◽  
Subunit Gene

Characterisation of three shoot apical meristem mutants of Arabidopsis thaliana

Development ◽  
1992 ◽  
Vol 116 (2) ◽  
pp. 397-403 ◽  
Author(s):  
H. M. Ottoline Leyser ◽  
I. J. Furner
Keyword(s):  
Arabidopsis Thaliana ◽  
Root Growth ◽  
Shoot Apical Meristem ◽  
Apical Meristem ◽  
Floral Development ◽  
Wild Type ◽  
Recessive Mutations ◽  
Phenotypic Characterisation ◽  
Dicotyledonous Plants ◽  
And Function

The shoot apical meristem of dicotyledonous plants is highly regulated both structurally and functionally, but little is known about the mechanisms involved in this regulation. Here we describe the genetic and phenotypic characterisation of recessive mutations at three loci of Arabidopsis thaliana in which meristem structure and function are disrupted. The loci are Clavata1 (Clv1), Fasciata1 (Fas1) and Fasciata2 (Fas2). Plants mutant at these loci are fasciated having broad, flat stems and disrupted phyllotaxy. In all cases, the fasciations are associated with shoot apical meristem enlargement and altered floral development. While all the mutants share some phenotypic features they can be divided into two classes. The pleiotropic fas1 and fas2 mutants are unable to initiate wild- type organs, show major alterations in meristem structure and have reduced root growth. In contrast, clv1 mutant plants show near wild-type organ phenotypes, more subtle changes in shoot apical meristem structure and wild-type root growth.

Mercury weakens membrane anchoring of Na-K-ATPase

1992 ◽  
Vol 262 (5) ◽  
pp. F837-F842 ◽  
Author(s):  
E. Imesch ◽  
M. Moosmayer ◽  
B. M. Anner
Keyword(s):  
Atpase Activity ◽  
Model Compound ◽  
Beta Subunit ◽  
Alpha Subunit ◽  
Structural Effects ◽  
Subunit Interaction ◽  
Alpha Beta ◽  
Membrane Anchoring ◽  
Atpase Inhibition

The presence of circulating inhibitors able to decrease the renal Na-K-adenosinetriphosphatase (ATPase) activity (natriuretic hormones) was postulated some 30 years ago. In the present work, the natriuretic inhibitor HgCl2 was selected as a model compound for the structural characterization of a possible natriuretic pathway for Na-K-ATPase modification. The structural effects of Na-K-ATPase inhibition by HgCl2 were assessed by trypsinolysis of the blocked enzyme in comparison with untreated preparations. The results show that inactivation of Na-K-ATPase by HgCl2 leads to the release of the alpha-subunit from the membrane preferentially in the E2 conformation but also in the E1 conformation. Apparently, HgCl2 weakens the membrane anchoring of the alpha-subunit, presumably by loosening the alpha-beta-subunit interaction. By this mechanism, the sensitivity of the Na-K-ATPase to extracellular drugs, hormones, and antibodies, as well as to intracellular proteases and other regulatory factors, could be altered.

scholarly journals Mutations That Reduce Sinapoylmalate Accumulation in Arabidopsis thaliana Define Loci With Diverse Roles in Phenylpropanoid Metabolism

Genetics ◽  
2001 ◽  
Vol 159 (4) ◽  
pp. 1741-1749
Author(s):  
Max Ruegger ◽  
Clint Chapple
Keyword(s):  
Arabidopsis Thaliana ◽  
Secondary Metabolites ◽  
Plant Development ◽  
Phenylpropanoid Metabolism ◽  
Normal Plant ◽  
Wild Type ◽  
Developmentally Regulated ◽  
Recessive Mutations ◽  
Uv Illumination ◽  
Cell Specificity

Abstract The products of phenylpropanoid metabolism in Arabidopsis include the three fluorescent sinapate esters sinapoylglucose, sinapoylmalate, and sinapoylcholine. The sinapoylmalate that accumulates in cotyledons and leaves causes these organs to appear blue-green under ultraviolet (UV) illumination. To find novel genes acting in phenylpropanoid metabolism, Arabidopsis seedlings were screened under UV for altered fluorescence phenotypes caused by changes in sinapoylmalate content. This screen identified recessive mutations at four Reduced Epidermal Fluorescence (REF) loci that reduced leaf sinapoylmalate content. Further analyses showed that the ref mutations affected other aspects of phenylpropanoid metabolism and some led to perturbations in normal plant development. A second class of mutations at the Bright Trichomes 1 (BRT1) locus leads to modest reductions in sinapate ester content; however, the most notable phenotype of brt1 mutants is the development of hyperfluorescent trichomes that appear to contain elevated levels of sinapate esters when compared to the wild type. These results indicate that at least five new loci affecting the developmentally regulated accumulation of phenylpropanoid secondary metabolites in Arabidopsis, and the cell specificity of their distribution, have been identified by screening for altered UV fluorescence phenotypes.

Characterization of mutant alleles of myospheroid, the gene encoding the beta subunit of the Drosophila PS integrins.

Genetics ◽  
1992 ◽  
Vol 132 (2) ◽  
pp. 519-528 ◽  
Author(s):  
T A Bunch ◽  
R Salatino ◽  
M C Engelsgjerd ◽  
L Mukai ◽  
R F West ◽  
...  
Keyword(s):  
Chromosome Pairing ◽  
Protein Product ◽  
Beta Subunit ◽  
Wild Type ◽  
Muscle Attachment ◽  
Negative Interaction ◽  
Gene Encoding ◽  
Attachment Sites ◽  
Muscle Attachment Sites

Abstract This paper presents the characterization of nine alleles of myospheroid, which encodes the beta PS subunit of the Drosophila PS integrins. On Southern blots, the mysXB87, mysXN101 and mysXR04 genes yield restriction digest patterns similar to that seen for wild-type chromosomes, however the mys1 and mysXG43 genes contain detectable deletions. mys1, mysXB87 and mysXG43 make little or no stable protein product, and genetically behave as strong lethal alleles. For the mysXN101 mutation, protein product is seen on immunoblots and a reduced amount of beta PS protein is seen at muscle attachment sites of embryos; this mutant protein retains some wild-type function, as revealed by complementation tests with weak alleles. Protein is also seen on immunoblots from mysXR04 embryos, and this allele behaves as an antimorph, being more deleterious in some crosses than the complete deficiency for the locus. mysts2 and mysnj42 are typically lethal in various combinations with other alleles at high temperatures only, but even at high physiological temperatures, neither appears to eliminate gene function completely. The complementation behaviors of mysts1 and mysts3 are quite unusual and suggest that these mutations involve regulatory phenomena. For mysts3, the data are most easily explained by postulating transvection effects at the locus. The results for mysts1 are less straightforward, but point to the possibility of a chromosome pairing-dependent negative interaction.

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