Development of Mammalian Serum Albumin Affinity Purification Media by Peptide Phage Display

Sato, Aaron K.; Sexton, Daniel J.; Morganelli, Lee A.; Cohen, Edward H.; Wu, Qi; Conley, Greg; Streltsova, Zoya; Lee, Stan W.; Devlin, Mary; DeOliveira, Daniel B.; Enright, Jasmin; Kent, Rachel B.; Wescott, Charles R.; Ransohoff, Tom C.; Ley, Arthur C.; Ladner, Robert C.

doi:10.1021/bp010181o

Cited by 89 publications

(64 citation statements)

References 37 publications

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“…Several prefractionation approaches employing chromatographic adsorbents and immunoaffinity methods have been used to remove albumin (8,18,19). Many LMW proteins are poorly recovered, however, as albumin is known to act as a carrier and transport protein within blood and therefore is likely to bind many species of interest such as peptide hormones, cytokines, and chemokines (10).…”

Section: Resultsmentioning

confidence: 99%

“…anti-human serum albumin antibody columns, protein A/G) have been developed to remove abundant proteins such as albumin and immunoglobulins from serum prior to mass spectrometric analysis (8,9). One of the fundamental oversights of serum protein depletion methodologies, however, is that many important low molecular weight (LMW) 1 proteins or peptides can be concomitantly removed by this sample preparation process as well.…”

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confidence: 99%

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Characterization of the Low Molecular Weight Human Serum Proteome

Tirumalai

Chan

Prieto

et al. 2003

Molecular & Cellular Proteomics

785

742

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Serum potentially carries an archive of important histological information whose determination could serve to improve early disease detection. The analysis of serum, however, is analytically challenging due to the high dynamic concentration range of constituent protein/peptide species, necessitating extensive fractionation prior to mass spectrometric analyses. The low molecular weight (LMW) serum proteome is that protein/peptide fraction from which high molecular weight proteins, such as albumin, immunoglobulins, transferrin, and lipoproteins, have been removed. This LMW fraction is made up of several classes of physiologically important proteins such as cytokines, chemokines, peptide hormones, as well as proteolytic fragments of larger proteins. Centrifugal ultrafiltration of serum was used to remove the large constituent proteins resulting in the enrichment of the LMW proteins/ peptides. Because albumin is known to bind and transport small molecules and peptides within the circulatory system, the centrifugal ultrafiltration was conducted under solvent conditions effecting the disruption of protein-protein interactions. The LMW serum proteome sample was digested with trypsin, fractionated by strong cation exchange chromatography, and analyzed by microcapillary reversed-phase liquid chromatography coupled on-line with electrospray ionization tandem mass spectrometry. Analysis of the tandem mass spectra resulted in the identification of over 340 human serum proteins; however, not a single peptide from serum albumin was observed. The large number of proteins identified demonstrates the efficacy of this method for the removal of large abundant proteins and the enrichment of the LMW serum proteome.

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Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

Characterization of the Low Molecular Weight Human Serum Proteome

Tirumalai

Chan

Prieto

et al. 2003

Molecular & Cellular Proteomics

785

742

View full text Add to dashboard Cite

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“…These libraries code for cyclic disulfide peptides with 9 -12 aa enclosed within the disulfide loop. The libraries were screened by using standard protocols (35). Briefly, 293c11 cells were incubated with the phage libraries, washed with buffer A [PBS (pH 6) and 10 mM EDTA] to remove nonbinding phage and treated with 66 M hIgG to elute the phage bound to FcRn.…”

Section: Transfection Of Hek293 Cells With Hfcrn For Screening and Bimentioning

confidence: 99%

Reduction of IgG in nonhuman primates by a peptide antagonist of the neonatal Fc receptor FcRn

Mezo

McDonnell

Hehir

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

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The neonatal Fc receptor FcRn provides IgG molecules with their characteristically long half-lives in vivo by protecting them from intracellular catabolism and then returning them to the extracellular space. Other investigators have demonstrated that mice lacking FcRn are protected from induction of various autoimmune diseases, presumably because of the accelerated catabolism of pathogenic IgGs in the animals. Therefore, targeting FcRn with a specific inhibitor may represent a unique approach for the treatment of autoimmune disease or other diseases where the reduction of pathogenic IgG will have a therapeutic benefit. Using phage display peptide libraries, we screened for ligands that bound to human FcRn (hFcRn) and discovered a consensus peptide sequence that binds to hFcRn and inhibits the binding of human IgG (hIgG) in vitro. Chemical optimization of the phage-identified sequences yielded the 26-amino acid peptide dimer SYN1436, which is capable of potent in vitro inhibition of the hIgG-hFcRn interaction. Administration of SYN1436 to mice transgenic for hFcRn induced an increase in the rate of catabolism of hIgG in a dose-dependent manner. Treatment of cynomolgus monkeys with SYN1436 led to a reduction of IgG by up to 80% without reducing serum albumin levels that also binds to FcRn. SYN1436 and related peptides thus represent a previously uncharacterized family of potential therapeutic agents for the treatment of humorally mediated autoimmune and other diseases.FcRn antagonist ͉ protein-protein interactions ͉ phage display ͉ autoimmune disease

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“…These are large enough to tolerate labeling, but small enough to be rapidly excreted and pass through endothelial cell fenestrae to extravascular tissues like small molecule drugs. Specific binding peptides can also be discovered de novo within large, diverse 'libraries' using phage display technology (5). The binding strength of de novo examples has, however, tended to be low like the smaller de novo antibody fragments (K d $ 100-1000 nM), requiring lengthy optimization campaigns.…”

Section: Heterodimersmentioning

confidence: 99%

Adventures in multivalency, the Harry S. Fischer memorial lecture CMR 2005; Evian, France

Tweedle¹

2006

Contrast Media Mol Imaging

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This review discusses multivalency in the context of drug discovery, specifically the discovery of new diagnostic imaging and related agents. The aim is to draw attention to the powerful role that multivalency plays throughout research involving molecular biology, in general, and much of biochemically targeted contrast agent research, in particular. Two examples from the author's laboratory are described. We created small ($5 kDa) peptide 'dimers' composed of two different, chemically linked peptides. The monomer peptides both bound to the same target protein with K d % 100 s nM, while the heterodimers had sub-nM K d values. Biological activity was evident in the heterodimers where none or very little existed in homodimers, monomers or monomer mixtures. Two different tyrosine kinases (KDR and C-Met) and four peptide families produced consistent results: multivalent heterodimers were uniquely different. The second example begins with making two micron ultrasound bubbles coated with the peptide, TKPPR (a Tuftsin antagonist) as a negative control for bubbles targeted with angiogenesis target-binding peptides. Unexpected binding of a 'negative' control, (TKPPR)-targeted bubble to endothelial cells expressing angiogenesis targets, led to the surprising result that TKPPR, only when multimerized, binds avidly, specifically and actively to neuropilin-1, a VEGF co-receptor. VEGF is the primary stimulator of angiogenesis. Tuftsin is a small peptide (TKPR) derived from IgG that binds to macrophages during inflammation, and has been studied for over 30 years. The receptor has never been cloned. The results led to new conclusions about Tuftsin, neuropilin-1 and the purpose, up to now unknown, of exon 8 in VEGF. Multivalency can be used rationally to solve practical problems in drug discovery. When targeting larger structures, multivalency is frequently unavoidable, and can lead to unpredictable and useful biochemical information, as well as to new drug candidates.

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Development of Mammalian Serum Albumin Affinity Purification Media by Peptide Phage Display

Cited by 89 publications

References 37 publications

Characterization of the Low Molecular Weight Human Serum Proteome

Characterization of the Low Molecular Weight Human Serum Proteome

Reduction of IgG in nonhuman primates by a peptide antagonist of the neonatal Fc receptor FcRn

Adventures in multivalency, the Harry S. Fischer memorial lecture CMR 2005; Evian, France

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