Selection of peptidic mimics of digoxin from phage-displayed peptide libraries by anti-digoxin antibodies 1 1Edited by I. A. Wilson

Ball, William J.; Zhenming, Wang; Malik, Bela; Kasturi, Rama; Dey, Purabi; Short, Mary K.; Margolies, Michael N.

doi:10.1006/jmbi.2000.3934

Cited by 7 publications

(8 citation statements)

References 43 publications

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“…The solution structure of the larger peptide, NPY (36 amino acids, molecular mass 4271 Da), with a length of 51 Å, 39 shows a sixturn α-helix at its C-terminus. Thus, AT is close to the size range of the 26-10 binding peptides (15mer around 1800 Da, 26mer around 2900 Da) isolated by Ball et al, 35 whereas NPY is much longer.…”

Section: Discussionmentioning

confidence: 62%

“…Additional similarities between 26-10 and antipeptide antibodies were observed in the SDR usage, indicating that the side chains at the putative SDRs of 26-10 are well positioned to interact with peptide antigens. 26-10 is also known to bind to peptides, 35 perhaps because of its anti-peptide like binding pocket topography and SDR side-chain orientation. Other desirable features of the 26-10 antibody as a scaffold for synthetic library construction include its high expression level in E. coli and its thermodynamic stability.…”

Section: Discussionmentioning

confidence: 98%

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Synthetic Antibody Libraries Focused Towards Peptide Ligands

Cobaugh

Almagro²,

Pogson

et al. 2008

Journal of Molecular Biology

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Synthetic antibody libraries have proven immensely useful for the de novo isolation of antibodies without the need for animal immunization. Recently, focused libraries designed to recognize particular classes of ligands, such as haptens or proteins, have been employed to facilitate the selection of high-affinity antibodies. Focused libraries are built using V regions encoding combinations of canonical structures that resemble the structural features of antibodies that bind the desired class of ligands and sequence diversity is introduced at residues typically involved in recognition. Here we describe the generation and experimental validation of two different single-chain antibody variable fragment libraries that efficiently generate binders to peptides, a class of molecules that has proven to be a difficult target for antibody generation. First, a human anti-peptide library was constructed by diversifying a scaffold: the human variable heavy chain (V(H)) germ line gene 3-23, which was fused to a variant of the human variable light chain (V(L)) germ line gene A27, in which L1 was modified to encode the canonical structure found in anti-peptide antibodies. The sequence diversity was introduced into 3-23 (V(H)) only, targeting for diversification residues commonly found in contact with protein and peptide antigens. Second, a murine library was generated using the antibody 26-10, which was initially isolated based on its affinity to the hapten digoxin, but also binds peptides and exhibits a canonical structure pattern typical of anti-peptide antibodies. Diversity was introduced in the V(H) only using the profile of amino acids found at positions that frequently contact peptide antigens. Both libraries yielded binders to two model peptides, angiotensin and neuropeptide Y, following screening by solution phage panning. The mouse library yielded antibodies with affinities below 20 nM to both targets, although only the V(H) had been subjected to diversification.

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Section: Discussionmentioning

confidence: 62%

Section: Discussionmentioning

confidence: 98%

Synthetic Antibody Libraries Focused Towards Peptide Ligands

Cobaugh

Almagro²,

Pogson

et al. 2008

Journal of Molecular Biology

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“…31 Such Oua-like immunoreactive compounds, therefore, are distinct from our isolates purified using mAb 1-10 because these latter contain both a Na ϩ pump inhibitor and Oua-binding proteins. It should be emphasized that the identity of the Na ϩ pump inhibitor in our current studies is unknown and may or may not be the same or related to OLC that we have previously isolated from bovine hypothalamus and characterized biochemically and physiologically in detail.…”

Section: Discussionmentioning

confidence: 82%

Ouabain-Binding Protein(s) From Human Plasma

et al. 2002

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Abstract-Conservation of the binding site on mammalian Na ϩ ,K ϩ -ATPase for cardiac glycosides and the importance of the Na ϩ pump in mammalian cellular physiology has stimulated the search for a mammalian analog of these plant compounds. One candidate, isolated from brain and blood, appears to be ouabain itself or a closely related isomer, the ouabain-like compound. Little is known about the circulating form. Because human steroid hormones circulate with carrier proteins, we produced a ouabain-specific monoclonal antibody (mAb 1-10) and used it to probe normal human plasma for ouabain-protein carrier complex. Ouabain-like biological activity was isolated in association with protein bands of 80, 50, and 25 kDa. These proteins appear to be human immunoglobulins or immunoglobulin-like because they are recognized by anti-human immunoglobulin antibodies, but not by anti-mouse immunoglobulin antibodies. The protein-containing fractions inhibit the binding of mAb 1-10 to immobilized ouabain, and with further purification on protein A, the immunoglobulin-like protein binds radioactive ouabain with an IC 50 of 200 to 600 nmol/L, but binds digoxin with 100-fold less affinity, suggesting specificity for ouabain or its isomer. Active protein fractions after purification on C 18 inhibit Na ϩ pump activity in human erythrocytes (IC 50Ϸ 4 nmol/L, ouabain equivalents), and this chromatography appears to dissociate the ouabain-like compound from the immunoglobulin protein(s). These immunoglobulin-like molecules may represent a subset of immunoglobulins (Յ0.5% of total protein A immunoglobulin) that function as a reservoir and delivery system for ouabain-like compounds in the modulation of human Na ϩ ,K ϩ -ATPase in vivo. (Hypertension. 2002;40:220-228.)

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“…Most scholars ascribed this mechanism to conformational structure mimicking (Ball et al, 2000;Yuan et al, 1999). In our research, the common amino sequences of those peptides mostly consisted of nonpolar amino acid residues and bore few similarities with OTA in the structure (Fig.…”

Section: Discussionmentioning

confidence: 92%

“…Phage-displayed short peptide libraries have been widely used (Romanov, 2003;Sidhu, Fairbrother, & Deshayes, 2003). They had been shown to be powerful tools for selecting peptide mimics of small-molecular-weight haptens including deoxynivalenol, aXatoxin B 1 , and digoxin (Ball et al, 2000;Thirumala-Devi, Miller, Reddy, Reddy, & Mayo, 2001;Yuan et al, 1999).…”

Section: Introductionmentioning

confidence: 97%

An immunoassay for ochratoxin A without the mycotoxin

Liu¹,

Yu²,

He³

2007

Food Control

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Selection of peptidic mimics of digoxin from phage-displayed peptide libraries by anti-digoxin antibodies 1 1Edited by I. A. Wilson

Cited by 7 publications

References 43 publications

Synthetic Antibody Libraries Focused Towards Peptide Ligands

Synthetic Antibody Libraries Focused Towards Peptide Ligands

Ouabain-Binding Protein(s) From Human Plasma

An immunoassay for ochratoxin A without the mycotoxin

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