ON THE DEVELOPMENT OF INCREASED TRYPTOPHAN SYNTHETASE ENZYME ACTIVITY BY CELL-FREE EXTRACTS OF NEUROSPORA CRASSA

1959 ◽  
Vol 37 (1) ◽  
pp. 1417-1430 ◽  
Author(s):  
S. D. Wainwright
Keyword(s):  
Energy Source ◽  
Neurospora Crassa ◽  
Calcium Ions ◽  
Polyvinyl Acetate ◽  
Enzymic Activity ◽  
Inorganic Salts ◽  
Synthetase Activity ◽  
Cell Free Extract ◽  
A Cell ◽  
Tryptophan Synthetase

A cell-free extract from conidia of Neurospora crassa developed increased tryptophan synthetase activity when incubated with a mixture of amino acids, a source of energy, and inorganic salts. Fifteen individual acids were required for development of maximal increases of enzymic activity. Either hexose diphosphate or adenosine triphosphate could serve as energy source. Diphosphopyridine nucleotide, polyvinyl acetate, and magnesium and calcium ions were required. The addition of ribonuclease enzyme, ethionine, fluorophenylalanine, thienylalanine, or various antibiotics prevented development of increased tryptophan synthetase activity. The increase of activity was accompanied by a net synthesis of protein.

ON THE DEVELOPMENT OF INCREASED TRYPTOPHAN SYNTHETASE ENZYME ACTIVITY BY CELL-FREE EXTRACTS OF NEUROSPORA CRASSA

1959 ◽  
Vol 37 (12) ◽  
pp. 1417-1430 ◽  
Author(s):  
S. D. Wainwright
Keyword(s):  
Energy Source ◽  
Neurospora Crassa ◽  
Calcium Ions ◽  
Polyvinyl Acetate ◽  
Enzymic Activity ◽  
Inorganic Salts ◽  
Synthetase Activity ◽  
Cell Free Extract ◽  
A Cell ◽  
Tryptophan Synthetase

A cell-free extract from conidia of Neurospora crassa developed increased tryptophan synthetase activity when incubated with a mixture of amino acids, a source of energy, and inorganic salts. Fifteen individual acids were required for development of maximal increases of enzymic activity. Either hexose diphosphate or adenosine triphosphate could serve as energy source. Diphosphopyridine nucleotide, polyvinyl acetate, and magnesium and calcium ions were required. The addition of ribonuclease enzyme, ethionine, fluorophenylalanine, thienylalanine, or various antibiotics prevented development of increased tryptophan synthetase activity. The increase of activity was accompanied by a net synthesis of protein.

A RIBONUCLEIC ACID FRACTION OF NEUROSPORA CRASSA EVOKING DEVELOPMENT OF A PSEUDO-TRYPTOPHAN SYNTHETASE ACTIVITY BY EXTRACTS OF MUTANTS TD1, TD2, AND TD3

1965 ◽  
Vol 43 (11) ◽  
pp. 1813-1828 ◽  
Author(s):  
S. D. Wainwright ◽  
E. Sandra McFarlane
Keyword(s):  
Neurospora Crassa ◽  
Type Strain ◽  
Wild Type Strain ◽  
Enzymic Activity ◽  
Synthetase Activity ◽  
Acid Fraction ◽  
Wild Type ◽  
Mutant Strains ◽  
Tryptophan Synthetase

"Soluble RNA" fractions isolated from the wild-type strain of Neurospora crassa evoked development of a pseudo-tryptophan synthetase enzyme activity in vitro by extracts of mutant strains lacking ability to produce the enzymic activity. The RNA fractions contained no detectable "template RNA". Some properties of the component eliciting development of pseudo-tryptophan synthetase activity are reported.

Isolation and partial characterization of amphibian tyrosine oxidase polysomes

Development ◽  
1970 ◽  
Vol 24 (1) ◽  
pp. 109-118
Author(s):  
E. L. Triplett ◽  
R. Herzog ◽  
L. P. Russell
Keyword(s):  
Amino Acids ◽  
Protein Synthesis ◽  
Amino Acid ◽  
Enzymic Activity ◽  
Antigenic Determinants ◽  
Free System ◽  
A Cell ◽  
Generating System ◽  
Soluble Components

A population of polysomes isolated from frogskinis capable of supporting protein synthesis in a cell-free system containing an energy generating system, ‘soluble components’, and amino acids. These polysomes catalyse the oxidation of DOPA after gentle trypsinization, and they also have antigenic determinants attributable to tyrosine oxidase. Skin polysomes sedimented in 10–30 % sucrose gradients contain tyrosine oxidase peaks of enzymic activity at the bottom and top of the tube and in the 250 S regions. A peak of tyrosine oxidase antigenic acitvity is found in the 250–350S region of the gradient. Polysomes resolved on the gradient retain the ability to support protein synthesis in a cellfree system. All 250–350S particles capable of supporting the incorporation of [14C]amino acid into tyrosine oxidase are precipitable with tyrosine oxidase antibodies. It is probable that 250–350S tyrosine oxidase antibody precipitates contain only polysomes for this protein.

Involvement of tyrosyl-tRNA synthetase in splicing of group I introns in Neurospora crassa mitochondria: biochemical and immunochemical analyses of splicing activity

1989 ◽  
Vol 9 (5) ◽  
pp. 2089-2104
Author(s):  
A L Majumder ◽  
R A Akins ◽  
J G Wilkinson ◽  
R L Kelley ◽  
A J Snook ◽  
...  
Keyword(s):  
Neurospora Crassa ◽  
Molecular Mass ◽  
Mitochondrial Protein ◽  
Gel Filtration ◽  
Nuclear Gene ◽  
Trna Synthetase ◽  
Synthetase Activity ◽  
Group I Introns ◽  
Group I

We reported previously that mitochondrial tyrosyl-tRNA synthetase, which is encoded by the nuclear gene cyt-18 in Neurospora crassa, functions in splicing several group I introns in N. crassa mitochondria (R. A. Akins and A. M. Lambowitz, Cell 50:331-345, 1987). Two mutants in the cyt-18 gene (cyt-18-1 and cyt-18-2) are defective in both mitochondrial protein synthesis and splicing, and an activity that splices the mitochondrial large rRNA intron copurifies with a component of mitochondrial tyrosyl-tRNA synthetase. Here, we used antibodies against different trpE-cyt-18 fusion proteins to identify the cyt-18 gene product as a basic protein having an apparent molecular mass of 67 kilodaltons (kDa). Both the cyt-18-1 and cyt-18-2 mutants contain relatively high amounts of inactive cyt-18 protein detected immunochemically. Biochemical experiments show that the 67-kDa cyt-18 protein copurifies with splicing and synthetase activity through a number of different column chromatographic procedures. Some fractions having splicing activity contain only one or two prominent polypeptide bands, and the cyt-18 protein is among the few, if not only, major bands in common between the different fractions that have splicing activity. Phosphocellulose columns resolve three different forms or complexes of the cyt-18 protein that have splicing or synthetase activity or both. Gel filtration experiments show that splicing activity has a relatively small molecular mass (peak at 150 kDa with activity trailing to lower molecular masses) and could correspond simply to dimers or monomers, or both, of the cyt-18 protein. Finally, antibodies against different segments of the cyt-18 protein inhibit splicing of the large rRNA intron in vitro. Our results indicate that both splicing and tyrosyl-tRNA synthetase activity are associated with the same 67-kDa protein encoded by the cyt-18 gene. This protein is a key constituent of splicing activity; it functions directly in splicing, and few, if any, additional components are required for splicing the large rRNA intron.

scholarly journals Hyaluronate synthetase inhibition by normal and transformed human fibroblasts during growth reduction

1987 ◽  
Vol 104 (4) ◽  
pp. 1105-1115 ◽  
Author(s):  
K Matuoka ◽  
M Namba ◽  
Y Mitsui
Keyword(s):  
Cell Proliferation ◽  
Cell Density ◽  
Human Fibroblasts ◽  
Transformed Cells ◽  
Synthetase Activity ◽  
Intact Cells ◽  
Free System ◽  
Glycosaminoglycan Synthesis ◽  
Cell Free System ◽  
A Cell

To establish the relation of glycosaminoglycan synthesis to cell proliferation, we investigated the synthesis of individual glycosaminoglycan species by intact cells and in a cell-free system, using normal and transformed human fibroblasts under differing culture conditions. Reducing serum concentration brought about a marked decline in the synthesis of hyaluronate (HA) as well as cell proliferation on both normal and transformed cells. Both HA synthesis and proliferation decreased with increasing cell densities markedly (in inverse proportion to cell density) in normal cells but gradually in transformed cells. This noticeable congruity of the changes in HA synthesis and proliferation indicates that the change in HA synthesis is related primarily to cell proliferation rather than to cell density or cellular transformation. Examination of HA synthesis in a cell-free system demonstrated that the activity of HA synthetase also fluctuated in conjunction with cell proliferation. Furthermore, growth-reduced cells (except crowded transformed cells) inhibited cell-free HA synthesis and this inhibition was induced coincidentally with a decrease in both HA synthetase activity and proliferation. These findings suggest that the change in HA synthesis is significant in the regulation of cell proliferation.

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