Recombinant Biomolecules

Mass Spectrometry in Sports Drug Testing Characterization of Prohibited Substances and Doping Control Analytical Assays, By Mario Thevis Copyright 2010 John Wiley Sons, Inc. 

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In several publications the mass spectral analysis of intact MAbs has been described.7475 MAbs are fairly big and complex, mostly recombinant biomolecules, constructed of two heavy-chain and two light-chain polypeptides that are covalently... 

Sayers JR. Acres of antibodies the future of recombinant biomolecule production Trends in Biotechnology 19 429—430 (2001). 

Recombinant DNA technology now verges on the ability to engineer at will the genetic constitution of organisms for desired ends. The commercial production of therapeutic biomolecules in microbial cultures is already established (for example, the production of human insulin in quantity in E. coli cells). Agricultural crops with desired attributes, such as enhanced resistance to her-... 

What importance could vinylphosphonic acid have for the synthesis of important biomolecules Its photolysis gives many oxidized products, including phosphoac-etaldehyde. This analogue of glycol aldehyde phosphate seems to be of interest its formation involves the recombination of hydroxyl radicals with vinylphosphonic acid radicals. 

Many of the methods developed to study protein interactions use the bait/prey model to detect interacting partners (Phizicky and Fields, 1995 Archakov et al., 2003 Piehler, 2005). The bait protein is a purified protein (often recombinant) that is used to lure and capture a putative interacting protein or biomolecule. The bait protein may be immobilized to a solid phase for affinity separations or be used in solution. It also may be fusion tagged (i.e., GST or 6X His) or labeled with a detectable molecule, such as a fluorescent probe. It often is the case... 

Fenn JB, Mann M, Meng CK Electrospray ionization for mass spectrometry of large biomolecules. Science (1989) 246 64-71. Patrick JS, Lagu AL Review applications of capillary electrophoresis to the analysis of biotechnology-derived therapeutic proteins. Electrophoresis (2001) 22 4179-4196. Sowell J, Salon J, Strekowski L, et al Covalent and noncovalent labeling schemes for near-infrared dyes in capillary electrophoresis protein applications. Methods Mol. Biol. (2004) 276 39-75. Moini M Capillary electrophoresis mass spectrometry and its application to the analysis ofbiological mixtures. Anal. Bio-anal. Chem. (2002) 373 466 180. Nemunaitis J, Holmlund JT, Kraynak M, et al. Phase I evaluation of ISIS 3521, an antisense oligodeoxynucleotide to protein kinase C-a, in patients with advanced cancer./. Clin. Oncol. (1999) 17 3586-3595. De Frutos M, Cifuentes A, Diez-Masa JC Differences in capillary electrophoresis profiles of urinary and recombinant erythropoietin. Electrophoresis (2003) 24 678-680. 

The advent of recombinant DNA technology has led to an increased interest in the structural characterization of proteins by spectroscopic methods. Few spectroscopic techniques can provide the amount of information regarding protein secondary and tertiary structure which can be obtained from circular dichroism (CD) spectroscopy. In this chapter we describe the capabilities of CD spectroscopy to provide details on the globular structure of proteins. In addition, we will provide an overview of quantitative secondary structure estimates via CD spectroscopy and of specialized CD methods for studying proteins in contact with membranes and other biomolecules. Certain aspects of protein CD spectroscopy have been previously reviewed [1-19]. 

This mode of separation, as the name suggests, uses stationary phases with a special affinity for a specific analyte. The affinity ligand immobilized on the stationary phase varies dramatically from peptide, to protein, to oligonucleotide, to monoclonal antibody. In some cases the target molecule is labelled with an affinity tag to simplify the separation. This approach is common in the synthesis of recombinant proteins where the system can be engineered so that the target biomolecule expresses a tag such as polyhistidine. A stationary phase functionalized with aminodiacetic acid and nickel chelate is then used to fish out the required molecule by chelating with the polyhistidine tag. 

The ProLabel peptide can be chemically conjugated or recombinantly fused to various biomolecules. To probe molecular interactions, EEC assays are based on a competition between the free and the ProLabel peptide-conjugated form of the biomolecule involved in the interaction under study. Bound to its interaction partner, the ProLabel-labelled biomolecule is not able to complement with the Enzyme Acceptor. Therefore the signal generated by the P-galactosidase is proportional to the concentration of the free biomolecule in the assay."... 

Microencapsulation has also been used medically for the encapsulation of live cells and vaccines. Biocompatibility can be improved by the encapsulation of artificial cells and biomolecules such as peptides, proteins, and hormones, which can prevent unwanted immunological reactions that would lead to inactivation or rejection. Microspheres are used for isolating materials until their activity is needed. The biotechnology industry employs microspheres to contain organisms and their recombinant products to aid in the isolation of these products. ... 

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