Peptide nucleic acid (PNA). A DNA mimic with a peptide backbone.
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DNA↗
Biomedical subjects
O Buchardt
Publications and source records attributed to O Buchardt.
Pitfalls in characterization of protein interactions using radioiodinated crosslinking reagents. Preparation and testing of a novel photochemical 125I-label transfer reagent.
Much attention has been focused on the study of protein interactions with radioiodinated photo-crosslinking reagents, and pitfalls in using this methodology are discussed. A new photochemical and cleavable heterobifunctional crosslinking reagent, succinimidyl N-14-(2-hydroxybenzoyl)-N-11-(4-azidobenzoyl)-9-oxo-8,11,14-triaza -4,5- dithiatetradecanoate (SHAD) was prepared, and its potential as a label transfer reagent was tested in model systems. SHAD was radioiodinated, and the labeled reagent (125I-SHAD) was converted to an amide (125I-HADM, as a mimicry of conjugation to protein 1) and photolyzed. When compared to the widely used SASD reagent (sulfosuccinimidyl 2-[[(4-azidosalicyl)-amino]ethyl]-1,3- dithiopropionate, Pierce), SHAD has a number of decisive advantages. The amide of 125I-SASD (125I-ASDM) was generated and photolyzed, and it was found that at least 50% of the radioactivity is released from 125I-ASDM after 3 min of irradiation, whereas only approximately 10% is liberated from 125I-HADM under similar conditions. Furthermore, 125I-HADM was photolyzed in the presence of excess amine (mimicry of crosslinking to protein 2), and the product was cleaved by reduction (mimicry of label transfer). The transformations in the course of photolysis were monitored by UV spectroscopy and TLC analysis, and a high degree of reagent cleavage upon reduction was demonstrated. 125I-SHAD was used to crosslink Lys78-plasminogen and fibrin. 125I-SHAD was conjugated to Lys78-plasminogen in the dark. Fibrinogen and thrombin were added, and Lys78-plasminogen was crosslinked to the fibrin clot by exposure to light.(ABSTRACT TRUNCATED AT 250 WORDS)
Azides↗
Monoclonal antibodies to thioguanine: influence of coupling position on fine specificity.
Thioguanine derivatives with reactive ester groups at positions 6, 7, or 9 of the purine ring were synthesized and coupled to a protein carrier. The purified protein derivative of tuberculin was used as the carrier for immunizing bacillus Calmette-Guerin primed mice. This led to high antibody titers against the homologously coupled hapten, and spleen cells from the immunized mice were used to produce monoclonal antibodies against thioguanine. All monoclonal antibodies were selected for their ability to recognize free thioguanine and were analyzed for their fine specificity by inhibition experiments with a panel of thiopurine derivates. The specificity of the monoclonal antibodies showed a strong dependence on the coupling position of the thioguanine. Within each group of monoclonal antibodies, raised against one of the three different conjugates, there was a high degree of heterogeneity, with antibodies differing in their binding according to the substitution on the thioguanine analogues used in the inhibition experiments. This panel of antibodies may be used for quantitative assays of thiopurines and their metabolites in patients undergoing treatment with thioguanine, 6-mercaptopurine, and azathioprine.
Animals↗
Molecular imprinting approach for the recognition of adenine in aqueous medium and hydrolysis of adenosine 5'-triphosphate.
Polymers capable of recognizing adenine in aqueous medium were developed by the molecular imprinting technique using methacrylic acid and 4(5)-vinylimidazole as comonomers with ethylene glycol dimethacrylate as the cross-linking agent under different polymerization conditions. The affinity of these polymers for adenine and other nucleotide bases was compared. The association constant for the binding of adenine to the polymer is calculated to be 4.3 x 10(3)M(-1). Furthermore, binding of adenosine 5'-triphosphate (ATP) to the polymers was evaluated, and an enhanced binding compared to adenine was observed. The binding of ATP is pH dependent, with the maximum around pH 3. Results have been explained based on the hydrogen bonding and ionic interactions between ATP and the ligands on the polymer matrix. The catalytic effect of these polymers for the hydrolysis of ATP is briefly discussed.
Adenine↗
C-terminal amidation of calcitonin by carboxypeptidase Y catalyzed transpeptidation with a photocleavable nucleophile.
The C-terminal amidation of calcitonin represents an important technological problem. A method using a serine carboxypeptidase-catalyzed transpeptidation reaction in combination with photochemical cleavage to give the warranted peptide amide is described. The overall yield is higher than 95%.
Amides↗