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Results for “Photoaffinity Labels”

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At least 1,063 records · Page 59Linked to original sources
PubMed1986

Photoaffinity labelling of a 33-35,000 dalton protein in cardiac, skeletal and smooth muscle membranes using a new 125I-labelled 1,4-dihydropyridine calcium channel antagonist.

The binding sites for Ca2+ channel antagonists were probed using Bay P 8857 [2-iodoethyl isopropyl 1,4-dihydropyridine-2,6-dimethyl-4-(3-nitrophenyl)-pyridine-3,5-dicarbox ylate] that has been radiolabelled with 125I. This drug was shown to bind with high affinity to cardiac, smooth, and skeletal muscle membranes, with a KD approximately equal to 0.3 nM. A protein of molecular weight 33-35,000 daltons was specifically and irreversibly radiolabelled after irradiation of cardiac, skeletal and aortic smooth muscle membranes, incubated with the [125I]-Bay P 8857. The peptide labelled by 1,4-dihydropyridine binding therefore appears similar in size for cardiac, skeletal, and smooth muscle. This data suggests that of the three peptide subunits which reportedly comprise the skeletal and cardiac muscle 1,4-dihydropyridine receptor complex, the 33-35,000 dalton peptide contains the dihydropyridine binding site.

Affinity Labels↗
PubMed1986

Long-lasting blockade of P2-receptors of the urinary bladder in vivo following photolysis of arylazido aminopropionyl ATP, a photoaffinity label.

In the present study the possibility of provoking an irreversible blockade of P2-receptors in vivo by photolyzing ANAPP3 was investigated. ANAPP3 administered as an intraarterial infusion produced a short-lived blockade of P2-receptor mediated contractions of cat urinary bladders in situ. However, irradiation of the bladders with visible light during the infusion of ANAPP3 resulted in a blockade of the response for at least 120 minutes. Thus, photoactivation of ANAPP3 in vivo produced an essentially irreversible blockade of P2-purinergic receptors.

Adenosine Triphosphate↗
PubMed1981

Induction of petite "mutants" in an ethidium-resistant strain of Saccharomyces cerevisiae by photoaffinity labeling. Distinction between early and late steps.

A strain of Saccharomyces cerevisiae (MH41-7B/011) was resistant to petite induction by ethidium bromide at 30 degrees, but was sensitive to induction by photolabeling with ethidium monoazide. These results suggested a defect in the mutant in metabolic activation of ethidium to account for its resistance. Synchronized cultures of both the mutant and the normal parent strains showed a substantial reduction in petite response to photolabeling in stationary phase cells which could not be accounted for by changes in cell penetration of the drug. The use of photolabeling with normal and mutant cells suggested that petite induction can be divided into early and late steps.

Drug Resistance, Microbial↗
PubMed1984

Production of frameshift mutations in Salmonella by phenanthridinium derivatives: enzymatic activation and photoaffinity labeling.

The effect of metabolic activation on the mutagenic potential of some phenanthridinium compounds was examined in Salmonella typhimurium strains TA1538 and TA1978 . All of the compounds tested were mutagenic in TA1538, a DNA excision-repair-deficient strain, when metabolizing enzymes were included in the assay. Reversions were not detected when these compounds were examined under the same conditions in TA1978 , the isogenic strain of TA1538 proficient in DNA repair. The mutagenic activity of an azido analog of propidium iodide was also examined using photoactivation and enzymatic activation, and with both conditions, reversions were observed in TA1538 but not in TA1978 . Furthermore, the ranking of mutagenic activity of propidium azide relative to ethidium azide analogs was comparable for both types of activation. The evidence from several studies suggests that the structural requirements for mutagenic activity for this series of phenanthridinium compounds appear to be the same whether mutagenesis is induced via photoactivation or metabolic activation. The interaction with DNA resulting in covalent alteration of the DNA is implicated as the mutagenic mechanism whether the active species is generated by metabolic- or photo-activation.

Affinity Labels↗
PubMed1992

Nonsteroidal estrogens bearing acyl azide functions: potential electrophilic and photoaffinity labeling agents for the estrogen receptor.

In an effort to develop novel affinity labeling agents for the estrogen receptor, we have synthesized two nonsteroidal ligands, a 1-aroyl-2-aryl tetralin system (1) and a 2-aryl-3-aroylbenzo[b]thiophene system (2). These agents, patterned after the Lilly antiestrogens trioxifene and LY 117018, respectively, embody acyl azide functions as part of a benzoyl chromophore. The acyl azide group has weak acylating activity, suitable for electrophilic affinity labeling, but this function is also photoreactive and, in its particular embodiment within these ligands, it could provide an efficient photochemical route to the highly reactive singlet acyl nitrene. The tetralin system (1) was prepared in nine steps from 6-methoxy-1-tetralone, and the benzothiophene system (2) was prepared in four steps from a known substituted benzo[b]thiophene precursor. In competitive binding assays, both compounds show reasonable binding affinity for the rat and lamb uterine estrogen receptor: estradiol = 100%, 1 = 3%, and 2 = 12%. When assayed by indirect receptor consumption assays, both compounds appear to have substantial capacity for irreversible binding (electrophilic reaction) with the receptor. This reactivity, which suggests that acylation of the receptor has occurred, is photoreversible. The nature of this ligand-receptor interaction is being investigated further.

Affinity Labels↗
PubMed1994

Photoaffinity labeling of rat liver microsomal steroid 5 alpha-reductase by 2-azido-NADP+.

Preincubation of female rat liver microsomal preparations with [2'-32P]2N3-NADP+ followed by photolysis with UV light (254 nm) and analysis by SDS-PAGE/autoradiography showed incorporation of 32P into at least 3 major protein bands in the molecular weight range of 14-97 Kd. Labeling of a 26 kD band, the apparent molecular weight of 5 alpha-reductase in liver microsomes, was accompanied by a loss of enzyme activity, consistent with its covalent modification. The inclusion of 20-fold excess NADP+ (100 microM) completely inhibited the incorporation of [2'-32P]2N3-NADP+ and preserved the enzyme activity, whereas excess NAD+ (100 microM) failed to protect 5 alpha-reductase (5 alpha R) activity. Similar results were obtained with the detergent-solubilized form of 5 alpha R. Polyethylene glycol (PEG) fractionation of detergent-solubilized preparations of 5 alpha R showed that all the 5 alpha R activity could be recovered in the 6.5% pellet with a 3-4-fold increase in the specific activity. photolysis of this fraction with [2'-32P]2N3-NADP+ resulted in approximately 2-fold increase in 32P labeling of the 5 alpha R band. Increasing photolysis time and concentration of the [2'-32P]2N3-NADP+ indicated that the half-life for photoincorporation and the apparent Kd were 1.0 min and 2 microM, respectively. These results suggest that 2N3-NADP+ is an effective probe of the NADP(H) binding site of 5 alpha R, and is a useful marker during purification of the enzyme.

3-Oxo-5-alpha-Steroid 4-Dehydrogenase↗