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PMID: 23280336 Published · ppublish English Journal Article Research Support, N.I.H., Extramural

Generating conformation-specific synthetic antibodies to trap proteins in selected functional states.

Methods (San Diego, Calif.) ·Vol. 60 ·No. 1 ·2013-03-15 ·Pages 3-14

Paduch M, Koide A, Uysal S, Rizk SS, Koide S, Kossiakoff AA

Abstract

A set of phage display sorting strategies and validation methodologies are presented that are capable of producing high performance synthetic antibodies (sABs) with customized properties. Exquisite control of antigen and conditions during the phage display selection process can yield sABs that: (1) recognize conformational states, (2) target specific regions of the surface of a protein, (3) induce conformational changes, and (4) capture and stabilize multi-protein complexes. These unique capabilities open myriad opportunities to study complex macromolecular processes inaccessible to traditional affinity reagent technology. We present detailed protocols for de novo isolation of binders, as well as examples of downstream biophysical characterization. The methods described are generalizable and can be adapted to other in vitro direct evolution approaches based on yeast or mRNA display.

MeSH Terms
Antibodies/chemistry,metabolism Antibody Specificity Enzyme-Linked Immunosorbent Assay Molecular Conformation Peptide Library Proteins/chemistry,metabolism
Chemicals
Antibodies Peptide Library Proteins
Authors & Affiliations
6 authors, click to expand affiliations / ORCID
Paduch Marcin
Department of Biochemistry and Molecular Biology, The University of Chicago, Chicago, IL 60637, USA.
Koide Akiko
Uysal Serdar
Rizk Shahir S
Koide Shohei
Kossiakoff Anthony A
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Article Info
Journal
Methods (San Diego, Calif.)
Abbr.
Methods
ISSN
1095-9130
DOI
10.1016/j.ymeth.2012.12.010
Published
2013-03-15
Epub
2012-00-29
Pages
3-14
Language
English
Region
United States
NLM ID
9426302
PMCID
PMC3650108
Subset
IM
Grants
NHGRI NIH HHS · U54 HG006436 · United States
NIGMS NIH HHS · U01 GM094588 · United States
NIGMS NIH HHS · R01 GM072688 · United States
NIGMS NIH HHS · U54 GM087519 · United States
NHGRI NIH HHS · U54-HG006436 · United States
NIGMS NIH HHS · U54-GM087519 · United States
NIGMS NIH HHS · R01-GM072688 · United States
NIGMS NIH HHS · U54 GM074946 · United States
NIGMS NIH HHS · U01-GM094588 · United States
External Links
PubMed source DOI: 10.1016/j.ymeth.2012.12.010 PMC Full Text View journal details
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