Bioconjugate preparation

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. 

Stolowitz, M.L. (1997) Phenylboronic acid complexes for bioconjugate preparation. US Patent No. 5,594,111. 

Wang D, Rogach AL, Caruso F (2002) Semiconductor quantum dot-labeled microsphere bioconjugates prepared by stepwise self-assembly. Nano Lett 2 857-861... 

Goldman, E. R., I. L. Medintz, A. Hayhurst, G. P. Anderson, J. M. Mauro, B. L Iverson, G. Georgiou, and H. Mattoussi. Self-assembled luminescent CdSe-ZnS quantum dot bioconjugates prepared using engineered poly-histidine terminated proteins. Anal. Chim. Acta 534, 63-67 (2005). 

Tyrosine residues are underutilized targets for bioconjugate preparation. As it is displayed with intermediate frequency on protein surfaces, tyrosine can often be modified with greater selectivity than other residues. In contrast to charged amino acids, tyrosine residues are often partially buried in the surface of the proteins owing to the amphipathic nature of the phenolic group, Fig. 10.3-3(a-d). This close association with the topography of protein... 

Figure 1.11 Structures of organometallic Au(l)-peptide bioconjugates. Prepared by Metzler-Nolte and coworkers [63].

ATRP is an exceptional tool for the synthesis of polymer-bioconjugates via grafting from technique from biomolecules such as proteins and peptides and even from high-value small molecule dmg entities. These molecules can be convenientiy modified to form suitable ATRP initiators to produce polymer-bioconjugates. Representative examples for polymer-bioconjugates prepared by ATRP using various functional biological initiators are illustrated in Fig. 2.23 [78]. 

Li, M., et al. 2008. Responsive polymer-protein bioconjugates prepared by RAFT polymerization and copper-catalyzed azide-alkyne click chemistry. Macromolecular Rapid Communications 29(12-13) 1172-1176. 

The aqueous Diels-Alder reaction has also been used for bioconjugate studies. A Diels-Alder reaction of diene oligonucleotides with maleimide dieneophiles was used to prepare oligonucleotide conjugates in aqueous media under mild conditions (Eq. 12.39).102 A Diels-Alder-type cycloaddition of an electronically matched pair of saccharide-linked conjugated dienes and a dienophile-equipped protein was the... 

Sakaguchi N, Kojima C, Harada A, Kono K (2008) Preparation of pH-sensitive poly (glycidol) derivatives with varying hydrophobicities their ability to sensitize stable liposomes to pH. Bioconjug Chem 19 1040-1048... 

The following sections describe several examples of saccharide modification for the purpose of bioconjugation, the study of glycan function, to prepare immunogens, or to increase the water solubility of a modified molecule. 

Mixtures of phospholipids in aqueous solution will spontaneously associate to form liposomal structures. To prepare liposomes having morphologies useful for bioconjugate or delivery techniques, it is necessary to control this assemblage to create vesicles of the proper size and shape. Many methods are available to accomplish this goal, however all of them have at least several steps in common (1) dissolving the lipid mixture in organic solvent, (2) dispersion in an aqueous phase, and (3) fractionation to isolate the correct liposomal population. 

The following sections discuss the concept and use of the (strept)avidin-biotin interaction in bioconjugate techniques. Preparation of biotinylated molecules and (strept)avidin conjugates also are reviewed with suggested protocols. For a discussion of the major biotinylation reagents, see Chapter 11 and Chapter 18, Section 3. 

Brinkley, M. (1992) A brief survey of methods for preparing protein conjugates with dyes, haptens, and cross-linking reagents. Bioconjugate Ghem. 3, 2. 

Ghetie, V., Till, M.A., Ghetie, M.-A., Tucker, T., Porter, J., Patzer, E.J., Richardson, J.A., Uhr, J.W., and Vitetta, E.S. (1990) Preparation and characterization of conjugates of recombinant CD4 and deglyco-sylated ricin A chain using different crosslinkers. Bioconjugate Chem. 1, 24-31. 

Holmberg, A., and Meurling, F. (1993) Preparation of sulfhydrylhorane-dextran conjugates for boron neutron capture therapy. Bioconjugate Chem. 4, 570-573. 

Miron, T., and Wilchek, M. (1993) A simplified method for the preparation of succinimidyl carbonate polyethylene glycol for coupling to proteins. Bioconjugate Ghent. 4, 568-569. 

Morpurgo, M., Veronese, F.M., Kachensky, D., and Harris, J.M. (1996) Preparation and characterization of polyethylene glycol)vinyl sulfone. Bioconjugate Chem. 7, 363-368. 

Noguchi, A., Takahashi, T., Yamaguchi, T., Kitamura, K., Takakura, Y., Hashida, M., and Sezaki, H. (1992) Preparation and properties of the immunoconjugate composed of anti-human colon cancer monoclonal antibody and mitomycin C—Dextran conjugate. Bioconjugate Chem. 3, 132-137. 

Singh, V., Mavila, A.K., and Kar, S.K. (1993) Comparison of the cytotoxic effect of hormonotoxins prepared with the use of heterobifunctional cross-linking agents N-succinimidyl 3-(2-pyridyldithio)propionate and N-succinimidyl 6-[3-(2-pyridyldithio)propionamido]-hexanoate. Bioconjugate Chem. 4, 473M82. 

A significant aid in the preparation of the second edition was the tremendous resources now available on the Internet for searching references to virtually any subject or key word within the scientific literature. For this reason, adding endless references to each chapter probably only would increase the size of the book by hundreds of pages, but add very little real value. Far better is for the reader to make use of pertinent Internet databases to search for key words, structure names, or reagent acronyms which can provide lists of hundreds or even thousands of additional references or links regarding any bioconjugation technique of interest. 

Scheme 1. Preparation of rhenium bioconjugates using the diamido-dithiolate bifunctional chelator.

Scheme 4. Preparation of bioconjugates using an N2S4 ligand system.

---