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B S Cooperman

Publications and source records attributed to B S Cooperman.

At least 145 records · Page 8Linked to original sources
PubMed1978

Photoinduced affinity labeling of the Escherichia coli ribosome puromycin site.

The photoincorporation of puromycin into Escherichia coli ribosomes has been studied in detail. Incorporation into protein L23 as a function of puromycin concentration follows a simple saturation curve and is specifically blocked by structural and functional analogues of puromycin, thus demonstrating that such incorporation proceeds via an affinity labeling process. Incorporation into L23 becomes more specific as the light fluence is reduced, indicating that such incorporation takes place from a native rather than light-denatured puromycin site. L23 remains the major labeled protein using ribosomes prepared by several procedures, suggesting the conservative nature of the site. In addition evidence is presented for affinity labeling of S14 and of a site in the RNA fraction of the 50S particle. Specific incorporation appears to proceed with an anomalously high quantum yield. The detailed photochemical mechanism is not understood, although 8-alkylation of purine moiety has been excluded. Incorporation is largely inhibited in the presence of thiol reagents.

Affinity Labels↗
PubMed1977

Identification of binding sites on the E. coli ribosome by affinity labeling.

Both electrophilic and photolabile derivatives of several different types of ribosomal ligands have been used in affinity labeling studies on the Escherichia coli ribosome. These studies have resulted in the localization of the peptidyl transferase center within a region of the 50S subunit, and the localization of the mRNA binding site within one of two regions on the 30S particle. In addition, labeling data have been obtained for GTP and streptomycin affinity labels. The affinity labeling results are discussed along with the results of other studies, and procedures are suggested for improving the resolving power of the affinity labeling technique as applied to ribosomes.

Affinity Labels↗
PubMed1976

Synthesis, structure determination, spectral properties, and energy-linked spectral responses of the extrinsic probe oxonol V in membranes.

The direct synthesis of bis]3-phenyl-5-oxoisoxazol-4-yl]pentamethineoxonol, which is shown to be the fluorescent probe OX-V (formerly MC-V), is described. The emission lifetime (0.9 +/- 0.1 ns) and the spectral properties of this dye in a number of systems are presented as well as the relative polarizations associated with the transition moments of the observable electronic transitions. The structure of OX-V was determined using elemental analysis and infrared and 1H nuclear magnetic resonance (NMR) spectroscopy. The use of the contact shift reagent, Eu(fod)3-d27, greatly facilitated the interpretation of the NMR results. In aqueous media, the anionic form of OX-V is present virtually exclusively due to the low solubility of the neutral species; formation of the latter species occurs when ethanol or methanol solutions of OX-V are acidiied. Both neutral and anionic dye forms can be detected in chloroform-ethanol solvents. The fluorescence intensity from excitation of the neutral species is an order of magnitude weaker than that from excitation of the anionic form and may result from the formation of excited anions due to the loss of a proton by the neutral species in the excited state. Polarization results indicate that the visible absorption of the dye is due to a single electronic transition. OX-V has been employed as a probe primarily in beef heart submitochondrial particles, reconstituted ATPase vesicles,a nd pigeon heart mitochondria. The energy-linked spectral changes of the probe in these preparations are described and mechanisms proposed for the spectral effects.

Animals↗
PubMed1975

Photoincorporation of puromycin and N-(ethyl-2-diazomalonyl)puromycin into Escherichia coli ribosomes.

[3H]Puromycin and N-(ethyl-2-diazomalonyl)[3H]puromycin are incorporated into E. coli ribosomes on irradiation at 253.7 nm. Both compounds incorporate into both protein and nucleic acid. Two-dimensional gel electrophoresis of ribosomal protein shows that L23 is the major protein labeled by puromycin. Although incorporation is clearly a complex process, evidence is presented that L23 is labeled via an affinity labeling process, thus placing L23 at the aminoacyl-tRNA receptor (A) site. N-(ethyl-2-diazomalonyl)puromycin is a ribosomal ligand, as shown by its inhibition of two ribosomal assays, but it is not a good puromycin analog, and it is unclear whether its incorporation, which proceeds via both carbene-dependent and carbene-independent processes, results from affinity labeling.

Azo Compounds↗
PubMed1973

Specific photoaffinity labeling of the adenosine 3':5'-cyclic monophosphate receptor in intact ghosts from human erythrocytes.

[(3)H]N(6)-(Ethyl 2-diazomalonyl)-adenosine 3':5'-cyclic monophosphate is incorporated into intact ghosts from human erythrocytes on photolysis at 253.7 nm. Incorporation is blocked in the presence of adenosine 3':5'-cyclic monophosphate (cyclic AMP) and does not occur in the absence of irradiation. Sodium dodecyl sulfate disc gel electrophoresis of solubilized ghosts shows that one protein is labeled. The position of this protein on the gel corresponds exactly to that previously found. [J. Biol. Chem. 247, 8145 (1972)] for the endogeneous protein substrate of the endogenous, cyclic AMP-dependent, protein kinase.

Azo Compounds↗