scholarly journals Application of high-performance liquid chromatography to the purification and characterization of ribosomal protein L3 from trichodermin-resistant yeast mutants

1986 ◽  
Vol 237 (2) ◽  
pp. 421-426 ◽  
Author(s):  
G J Threadgill ◽  
R C Conrad ◽  
L M Changchien ◽  
M Cannon ◽  
G R Craven
Keyword(s):  
Ribosomal Protein ◽  
High Performance ◽  
Reverse Phase ◽  
Exchange Method ◽  
Two Dimensional Gel Electrophoresis ◽  
Ribosomal Subunits ◽  
Direct Demonstration ◽  
Ribosomal Protein L3 ◽  
Peptide Profiles ◽  
Yeast Mutants

A new h.p.l.c. cation-exchange method has been used to separate proteins from 60S ribosomal subunits prepared from strains of Saccharomyces cerevisiae sensitive or resistant to trichodermin. Ribosomal protein L3 was identified in column eluates by one-dimensional and two-dimensional gel electrophoresis and purified further by reverse-phase h.p.l.c. The protein was cleaved with CNBr and the products were analysed, again by reverse-phase h.p.l.c. A marked difference was observed in the peptide profiles between preparations from trichodermin-sensitive and trichodermin-resistant yeast strains. These results provide the first direct demonstration that, in yeast, mutationally induced resistance to trichodermin can alter the covalent structure of ribosomal protein L3. They convincingly demonstrate the potential of the experimental technique for the rapid and preparative separation of a selected yeast ribosomal protein and its subsequent characterization.

scholarly journals A rapid and preparative method for the separation of yeast ribosomal proteins by using high-performance liquid chromatography

1987 ◽  
Vol 244 (3) ◽  
pp. 523-532 ◽  
Author(s):  
G J Threadgill ◽  
R C Conrad ◽  
M Cannon ◽  
G R Craven
Keyword(s):  
Ribosomal Proteins ◽  
High Performance ◽  
Preparative Method ◽  
Reverse Phase ◽  
Yeast Saccharomyces Cerevisiae ◽  
Two Dimensional Gel Electrophoresis ◽  
Preparative Scale ◽  
Ribosomal Subunits ◽  
Single Protein ◽  
Yeast Ribosomes

Ribosomal proteins from the yeast Saccharomyces cerevisiae were separated, on a preparative scale, by ion-exchange h.p.l.c. Proteins from the small and large ribosomal subunits were resolved, respectively, into 33 and 23 peaks, and most of the proteins present in these peaks were identified by using one- and two-dimensional gel electrophoresis. Several of the peaks appeared to contain a single protein uncontaminated by other species. Ribosomal proteins were also separated by using reverse-phase h.p.l.c. Analysis of the peaks resolved indicated that the order of elution for the proteins of both ribosomal subunits is, in certain cases, different for each of the two h.p.l.c. techniques used. Thus a combination of the two chromatographic methods employed here has the potential to facilitate the rapid and preparative separation of each of the proteins present in yeast ribosomes.

scholarly journals Phosphorylated and other modified forms of eukaryotic ribosomal protein S3 analysed by two-dimensional gel electrophoresis

1980 ◽  
Vol 189 (2) ◽  
pp. 241-245 ◽  
Author(s):  
D P Leader
Keyword(s):  
Ribosomal Protein ◽  
Gel Electrophoresis ◽  
Two Dimensional ◽  
Baby Hamster Kidney ◽  
Two Dimensional Gel Electrophoresis ◽  
Proteolytic Fragment ◽  
Normal Cells ◽  
Ribosomal Subunits ◽  
Ribosomal Protein S3 ◽  
Modified Forms

Proteins were isolated from the 40S ribosomal subunits of baby-hamster kidney fibroblasts and subjected to two-dimensional gel electrophoresis. When the cells were pretreated with cyclic AMP or 2-deoxyglucose a more basic derivative of ribosomal protein S3 or S3a was often observed, apparently similar to that previously reported to occur early in liver generation. This derivative was not a dephosphorylated form of protein S3, which protein does not appear to be phosphorylated in normal cells; nor did it correspond to the proteolytic fragment, S3b. It appears to be an oxidation product of protein S3 or S3a, as it can be eliminated by thorough reduction of the ribosomal protein before electrophoresis. In contrast with previous results with Krebs II ascites cells, starvation of baby-hamster kidney fibroblasts of glucose did not cause extensive phosphorylation of ribosomal protein S3.

Preparation and Characterization of NH2-Terminal Fibrinogen Bβ Fragments from N-DSK of Human Fibrinogen

1984 ◽  
Vol 51 (01) ◽  
pp. 016-021 ◽  
Author(s):  
S Birken ◽  
G Agosto ◽  
B Lahiri ◽  
R Canfield
Keyword(s):  
High Performance ◽  
Gel Filtration ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Reverse Phase ◽  
Human Fibrinogen ◽  
Human Volunteers ◽  
Cnbr Cleavage ◽  
Two Peaks

SummaryIn order to investigate the early release of NH2-terminal plasmic fragments from the Bβ chain of fibrinogen, substantial quantities of Bβ 1-42 and Bβ 1-21 are required as immunogens, as radioimmunoassay standards and for infusion into human volunteers to determine the half-lives of these peptides. Towards this end methods that employ selective proteolytic cleavage of these fragments from fibrinogen have been developed. Both the N-DSK fragment, produced by CNBr cleavage of fibrinogen, and Bβ 1-118 were employed as substrates for plasmin with the finding of higher yields from N-DSK. Bβ 1-42 and Bβ 1-21 were purified by gel filtration and ion-exchange chromatography on SP-Sephadex using volatile buffers. When the purified preparation of Bβ 1-42 was chromatographed on reverse-phase high performance liquid chromatography, two peaks of identical amino acid composition were separated, presumably due either to pyroglutamate or to amide differences.

Click Chemistry Expedited Radiosynthesis: Sulfur [18F]fluoride Exchange of Aryl Fluorosulfates

2019 ◽  
Author(s):  
Qinheng Zheng ◽  
Hongtao Xu ◽  
Hua Wang ◽  
Wen-Ge Han Du ◽  
Nan Wang ◽  
...  
Keyword(s):  
Click Chemistry ◽  
High Performance ◽  
Isotopic Exchange ◽  
Tumor Imaging ◽  
Radiochemical Yield ◽  
Exchange Method ◽  
Molar Activity ◽  
Positron Emission ◽  
Sulfur Fluoride

The lack of simple, efficient [<sup>18</sup>F]fluorination processes and new target-specific organofluorine probes remains the major challenge of fluorine-18-based positron emission tomography (PET). We report here a fast isotopic exchange method for the radiosynthesis of aryl [<sup>18</sup>F]fluorosulfate based PET agents enabled by the emerging sulfur fluoride exchange (SuFEx) click chemistry. The method has been applied to the fully-automated <sup>18</sup>F-radiolabeling of twenty-five structurally diverse aryl fluorosulfates with excellent radiochemical yield (83–100%) and high molar activity (up to 281 GBq µmol<sup>–1</sup>) at room temperature in 30 seconds. The purification of radiotracers requires no time-consuming high-performance liquid chromatography (HPLC), but rather a simple cartridge filtration. The utility of aryl [<sup>18</sup>F]fluorosulfate is demonstrated by the <i>in vivo</i> tumor imaging by targeting poly(ADP-ribose) polymerase 1 (PARP1).

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