Carbodiimide moieties

Modification of the ATPase by ATP-EDC occurred without significant incorporation of the nucleotide into the enzyme, suggesting that the inhibition of enzymatic activity is entirely due to intramolecular crosslinking by the carbodiimide moiety... 

As mentioned in the introduction, polymer-supported reagents can be used in excess to drive a reaction to completion, without a penalty in terms of purification. However, in many coupling reactions, it is not only an excess of coupling reagent, e.g., a carbodiimide moiety such as 10 (Scheme 2), that is required to drive the reaction to completion, but also an excess of one of the coupling partners. Consequently, new methods have been developed to separate these excess quantities from the product by simple filtration processes (Fig. 3). These techniques are described in more detail in Chapter 1. 

In the case of jV,jV-dialkyl-5-phenyl-6f/-l, 3,4-thiadiazin-2-amines, the reaction stops at the stage of 1 as ring opening and formation of a carbodiimide moiety are not possible wnth the 2-(Ar,A -dialkylamino) compounds. 

Amino-l,2,4-oxadiazoles 7 are obtained by reaction of amidoximes with 2 equiv. of carbodiimides in an analogous two-step process the second carbodiimide moiety is thought to accelerate amine elimination by trapping after ring-closure [464]. 

COOH groups of the PE (PAA or PGA) and the -NH2 moieties of the pre-loaded protein, using l-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) as a catalyst [99]. Negligible enzyme desorption (<0.1 %) is observed from cross-linked lysozyme-loaded MS spheres after exposing the samples to an aqueous solution for 48 h, while about 25 % of the immobilized lysozyme is desorbed under the same conditions when the lysozyme is not cross-linked. 

Figure 8 Chemiluminescent (A and B) and bioluminescent (C) detections for immobilized hybridizations of PCR product. Dg, digoxigenin Bt, biotin Ad, avidin. Procedure A [30] Biotin moiety is incorporated into PCR products during the amplification reaction, using one 5 -biotinylated primer. The product is hybridized with a Dg-labeled probe and is immobilized on streptavidin-coated magnetic beads. This capture reaction is carried out for 30 min at 37°C. A permanent magnet is used to sediment the beads during washing to remove unbound DNA. By incubation with the washed beads for 45 min at 37°C, anti-Dg antibody conjugated to HRP enzyme is bound to the Dg-labeled probe, and luminol reaction is performed for CL detection. Procedure B [31] Wells of apolystyrene microtiter plate are activated with l-ethyl-3-(3-dimethylaminopropyl)-carbodiimide, and then coated with a labeled cDNA probe complementary to an internal region of the target DNA.

FIGURE 2.27 More on additives. In a carbodiimide-mediated reaction between acid 1 and amine 2, addition of HOObt can lead to the side reaction of aminolysis at the carbonyl of the activating moiety of ester 3, generating addition product 4. Addition of HOBt to a mixed-anhydride reaction containing unconsumed chloroformate generates mixed carbonate 5, leading to production of urethane 6. 

Many types of mono-, bi-, and multifunctional coupling reagents are available for labeling antibodies or antigens with an enzyme. Glutaraldehyde, carbodiimide, N-succinimidyl-3-(2-pyridyldithio)propionate, and periodate oxidation of carbohydrate moieties to form active dialdehydes are several commonly used approaches in the preparation of enzyme conjugates (104-106). 

In general, most of published immunochemical methods on chloramphenicol assay are based on antibodies particularly specific for the aromatic ring and the propanediol moiety of chloramphenicol. This is due to the fact that the main chloramphenicol analogues such as thiamphenicol differ from chloramphenicol in one of these parts of the chloramphenicol molecule. To obtain such antibodies, chloramphenicol was linked to the carrier bovine serum albumin at the acyl chain of the molecule by mixed anhydride or carbodiimide reactions (49, 50). In both... 

Unfortunately, A-(9-fluorenylmethoxycarbonyl)aziridine-2-carboxylic acid cannot be used in peptide synthesis, since N-deprotection of the respective peptides with secondary amines leads to oxazoline or dehydroamino acid side products. Similarly, N-(tert-butoxy-carbonyl)aziridine-2-carboxylic acid is inappropriate due to the instability of the aziridine moiety to TFA treatment. Attempts to convert A-tritylaziridine-2-carboxylic acid into homogenous and stable active esters as useful intermediates in peptide synthesis leads to positive results only in the case of the pentafluorophenyl ester. 47 Consequently, this active ester seems to be the method of choice for acylating peptides. The related Abhydroxysuc-cinimide and A-3-hydroxy-4-oxo-3,4-dihydro-l,2,3-benzotriazine ester could not be isolated in pure form and have therefore been used as crude products. 47 Access to 2-carbonylazir-idine peptides is also possible by carbodiimide-mediated coupling. Additionally, alkylamides of A-tritylaziridine-2-carboxylic acid are prepared by the azide method,1 5 yet this method fails in peptide coupling steps. 85 ... 

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