Formation bioconjugates

The use of molecular biology methods, described in Section 5.3 seems to be especially worthwhile as it offers novel possibilities of optimization on process adjustment. Directed evolution leads to the formation of new biocatalysts with improved characteristics (selectivity, activity, stability, etc.). Incorporation ofnon-proteinogenic amino acids makes it possible to reach beyond the repertoire of building blocks used by nature. The prospect of bioconjugate preparation offers the possibility to form functional clusters of enzymes and to perform multiple synthetic steps in one pot. 

Ultimately, this section is meant to function as a ready-reference database for learning or review of bioconjugate chemistry. In this regard, a reaction can be quickly found, a short discussion of its properties and use read, and a visual representation of the chemistry of bond formation illustrated. What this section is not meant to be is an exhaustive discussion on the theory or mechanism behind each reaction, nor a review of every application in which each chemical reaction has been used. For particular applications where the chemistries are employed, cross-references are given to other sections in this book or to outside literature sources. 

Figure 15.8 The Bingel reaction for the modification of fullerenes involves the in situ formation of a reactive halogen species in the presence of the strong base DBU. The cyclopropanation product can be used to create many bioconjugates.

Nakajima, N., and Ikada, Y. (1995) Mechanism of amid formation by carbodiimides for bioconjugation in aqueous media. Bioconjugate Chem. 6, 123-130. 

Dithiolate ligands form stable anionic square pyramidal complexes with the Re03+ core [10]. This has been exploited in the dimer-captosuccinic acid complexes of 186Re and 188Re (vide infra), and in the use of the cyclic anhydride of dimercaptosuccinic acid as a bifunctional chelator for bioconjugate formation. The anhydride, in protected form such as 38, is reacted with antibody or other protein to form an amide or... 

An interesting approach to rhenium bioconjugate formation that has been developed but apparently has yet to be fully evaluated from the standpoint of applicability and stability is the use of chelating and nonchelating phosphine imine and phosphine oxide ligands [110]. For... 

Hong S, Leroueil PR, Janus EK, et al. (2006). Interaction of polycationic polymers with supported lipid bilayers and cells nanoscale hole formation and enhanced membrane permeability. Bioconjug. Chem. 17 728-34. 

M. Funk, I. Ponten, A. Seidel, B. Jemstrom, Critical Parameters for Adduct Formation of the Carcinogen (+)-a t/-Benzo[a]pyrene-7,8-dihydrodiol 9,10-Epoxide with Oligonucleotides , Bioconjugate Chem. 1997, 8, 310 - 317. 

Takeoka S, Sakai H, Kose T, et al. Methemoglobin formation in hemoglobin vesicles and reduction by encapsulated thiols. Bioconjug Chem 1997 8 539. 

Figure 14.4 Function and properties of a QD FRET-based nanosensor. Generalized QD bioconjugate nanosensor schematic. Each QD is surrounded by an average of —10—15 protein molecules. Formation of QD-protein-analogue assembly results in quenching of the QD emission. Adding preferred analyte to the solution displaces dye-labeled analogue from the sensor assembly, resulting in an increase in direct QD emission.

Addition of a nucleophile to the C-6 position of cytosine often results in fascile displacement reactions occurring at the N4 location. With hydroxylamine attack, nucleophilic displacement causes the formation of an N4-hydroxy derivative. A particularly important reaction for bioconjugate chemistry, however, is that of nucleophilic bisulfite addition to the C-6 position. Sulfonation of cytosine can lead to two distinct reaction products. At acid pH wherein the N-3 nitrogen is protonated, bisulfite reaction results in the 6-sulfonate product followed by spontaneous hydrolysis. Raising the pH to alkaline conditions causes effective formation of uracil. If bisulfite addition is done in the presence of a nucleophile, such as a primary amine or hydrazide compound, then transamination at the N4 position can take place instead of hydrolysis (Fig. 38). This is an important mechanism for adding spacer arm functionalities and other small molecules to cytosine-containing oligonucleotides (see Chapter 17, Section 2.1). 

Carbodiimide-mediated amide bond formation effectively occurs between pH 4.5 and 7.5. Buffer systems using MES or phosphate may be used to stabilize the pH during the course of the reaction. For additional information on specific carbodiimides used in bioconjugate chemistry, see Chapter 3, Section 1. 

R. Kircheis, and E. Wagner. 2001. Different strategies for formation of pegylated EGF-conjugated PEI/DNA complexes for targeted gene delivery. Bioconjug. Chem. 12 529-537. 

The oxidative introduction of carboxylic functions to nanotubes provides a large number of CNT-functional exploitations and permits covalent functionalization by the formation of amide and ester linkages and other carboxyl derivatives [24]. Bifunctional molecules (diamines, diols, etc.) are often utilized as linkers. More illustrative examples are nanotubes decorated with amino-functionalized dendrimers, nucleic acids, enzymes, etc., and the formation of bioconjugates of CNTs [96]. 

Ansell, S. M.,Tardi, P. G., and Buchkowsky, S. S. (1996), 3-(2-Pyridyldithio)propionic acid hydrazide as a cross-linker in the formation of liposome-antibody conjugates, Bioconjugate Chem.,1,490-496. 

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