Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Affinity Labels”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,063 records · Page 59Linked to original sources
PubMed1981

Affinity labeling of the ATP binding site of bovine lung cyclic GMP-dependent protein kinase with 5'-p-fluorosulfonylbenzoyladenosine.

Bovine lung cyclic GMP-dependent protein kinase was covalently labeled with the ATP analog, 5-p-fluorosulfonylbenzoyladenosine. The inactivation reaction was pseudo-first order. The rate of kinase sulfonation exhibited saturation kinetics indicative of a rapid reversible binding of the reagent prior to enzyme modification. The enzyme could be protected by MgATP, MgADP, and Mg-adenylylimidodiphosphate but not by a synthetic peptide substrate. Cyclic GMP when bound to the kinase did not influence the rate of labeling. The reagent demonstrated competitive inhibition with respect to MgATP; the Ki was found to be 0.82 mM. Magnesium and cobalt ions when included in the reaction mixture accelerated the inactivation rate up to severalfold. Addition of basic polypeptides such as mixed histone, protamine sulfate, or poly-L-lysine HBr also markedly accelerated the sulfonation rate. Inactivation of the kinase with 5-'fluorosulfonyl[3H]benzoyladenosine resulted in a linear relationship between the residual phosphotransferase activity and the incorporation of up to 0.9 mol of reagent/mol of monomer.

Adenosine↗
PubMed1982

Covalent affinity labeling by periodate-oxidized [alpha-32P]ATP of the large-T proteins of polyoma and SV40 viruses.

Incubation of crude extracts from cells lytically infected with polyoma virus in the presence of periodate-oxidized [alpha-32P]ATP led to the radioactive labeling of one main polypeptide immunoprecipitated by anti-T antigen antibodies. It was absent from extracts of mock-infected cells and exhibited an apparent Mr value of 105,000, identical with that of the large-T viral protein. This polypeptide was unambiguously identified as large-T on the basis of the heat sensitivity of the in vitro labeling in extracts from cells infected with a tsa mutant. The amount of incorporated radioactivity was found to increase linearly with that of infected cell extract and with the incubation time. Labeling resulted from the specific binding of an oxidized ATP molecule to a nucleotide binding site of the large-T protein, since it was efficiently completed by unlabeled ATP. Study of the dependence on the concentration of oxidized ATP evidenced a first order kinetic for the labeling reaction. Similar results were obtained for the large-T protein of SV40 virus.

Adenosine Triphosphate↗
PubMed1982

2-Bromoacetylaminopentitol 1,5-bisphosphate as an affinity label for ribulose bisphosphate carboxylase/oxygenase from Rhodospirillum rubrum.

2-Bromoacetylaminopentitol 1,5-bisphosphate (BrAcNH-pentitol-P2) (an epimeric mixture of 2-bromoacetylamino-2-deoxy-D-ribitol bisphosphate and 2-bromoacetylamino-2-deoxy-D-arabinitol 1,5-bisphosphate) has been synthesized from D-ribulose 1,5-bisphosphate by reductive amination with sodium cyanoborohydride followed by bromoacetylation of the resultant amine with bromoacetyl bromide. Under conditions that favor full activation of the enzyme, ribulose bisphosphate carboxylase/oxygenase from Rhodospirillum rubrum is completely inactivated by BrAcNH-pentitol-P2 in a pseudo-first order process. A rate saturation is observed with a minimal inactivation half-life of 38 min and Kinact for reagent of 0.38 mM. The competitive inhibitor 2-carboxyribitol 1,5-bisphosphate reduces the rate of inactivation, and kinetic analyses are consistent with the protection reflecting true competition of inhibitor and reagent for the same site. As shown with isotopically labeled reagent, complete inactivation is associated with covalent incorporation of 1.1 mol of reagent/mol of subunit. Based on reversibility of inactivation by thiolysis and based on analysis of labeled products in acid hydrolysates of the modified enzyme, a methionyl sulfonium salt is the reaction product. In the absence of CO2 and Mg2+ (ligands required for activation), the enzyme is resistant to BrAcNH-pentitol-P2, which suggests that the site-specific modification of a methionyl residue requires a fully functional catalytic center.

Amino Acids↗
PubMedDate not supplied

[Affinity labeling of ribosomes from Escherichia coli with 4-(N-2-chloroethyl, N-methylamino)-benzaldehyde actyl derivatives of oligouridylates].

4-(N-2-chloroethyl-N-methylamino)-benzaldehyde acetyl derivatives of penta-., hexa, hepta-, octauridylates were used for localization of the structures organizing the mRNA-binding centre of ribosomes. These derivatives, alike free oligonucleotides, stimulate the binding of phenylalanyl-tRNA to ribosomes. Within the specific complex all the oligonucleotide derivatives alkylated the 30S ribosomal subunit. Octauridylate and hexauridylate derivatives specifically alkylated also the 50S subunit of ribosomes. Polyuridylic acid protected ribosomal subunits from alkylation. In the 30S subunit the derivatives modify 16S RNA and proteins: S4, S5, S7, S9, S13, S15, S18, S21. It was found that oligouridylate derivatives of different length alkylate different proteins.

Escherichia coli↗