Locking the Elbow: Improved Antibody Fab Fragments as Chaperones for Structure Determination

Bailey LJ, Sheehy KM, Dominik PK, Liang WG, Rui H, Clark M, Jaskolowski M, Kim Y, Deneka D, Tang WJ, Kossiakoff AA

J. Mol. Biol. 2018 Feb;430(3):337-347

PMID: 29273204

Abstract

Antibody Fab fragments have been exploited with significant success to facilitate the structure determination of challenging macromolecules as crystallization chaperones and as molecular fiducial marks …

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Engineered synthetic antibodies as probes to quantify the energetic contributions of ligand binding to conformational changes in proteins

Mukherjee S, Griffin DH, Horn JR, Rizk SS, Nocula-Lugowska M, Malmqvist M, Kim SS, Kossiakoff AA

J. Biol. Chem. 2018 Feb;293(8):2815-2828

PMID: 29321208

Abstract

Conformational changes in proteins due to ligand binding are ubiquitous in biological processes and are integral to many biological systems. However, it is often challenging to link ligand-induced conformational changes to …

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Targeted rescue of cancer-associated IDH1 mutant activity using an engineered synthetic antibody

Rizk SS, Mukherjee S, Koide A, Koide S, Kossiakoff AA

Sci Rep 2017 Apr;7(1):556

PMID: 28373671

Abstract

We have utilized a high-diversity phage display library to engineer antibody fragments (Fabs) that can modulate the activity of the enzyme isocitrate dehydrogenase 1 (IDH1). We show that a conformation-specific Fab can reactivate an IDH1 mutant associated with …

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Generating Conformation and Complex-Specific Synthetic Antibodies

Paduch M, Kossiakoff AA

Methods Mol. Biol. 2017;1575:93-119

PMID: 28255876

Abstract

Phage display is commonly used to identify and isolate binders from large combinatorial libraries. Here we present phage selection protocols enabling generation of synthetic antibodies capable of recognizing multiprotein complexes and conformational states. The procedure describes stages of the experiment design, optimization, and screening, …

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Prolactin Receptor-Mediated Internalization of Imaging Agents Detects Epithelial Ovarian Cancer with Enhanced Sensitivity and Specificity

Sundaram KM, Zhang Y, Mitra AK, Kouadio JK, Gwin K, Kossiakoff AA, Roman BB, Lengyel E, Piccirilli JA

Cancer Res. 2017 Apr;77(7):1684-1696

PMID: 28202518

Abstract

Poor prognosis of ovarian cancer, the deadliest of the gynecologic malignancies, reflects major limitations associated with detection and diagnosis. Current methods lack high sensitivity to detect small tumors and high …

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Generation of Synthetic Antibody Fragments to Detergent Solubilized Membrane Proteins

Uysal S, Kossiakoff AA

Springer Protocols Handbook, p1-13

Abstract

Structural determination of membrane proteins is extremely challenging due to the physical characteristics of membrane proteins themselves and the lack of adequate tools and technologies to perform the studies. Recent developments in micro-focus X-ray beams, novel detergents, protein thermo-stabilization, and protein engineering have been essential in …

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The Preserved HTH-Docking Cleft of HIV-1 Integrase Is Functionally Critical

Galilee M, Britan-Rosich E, Griner SL, Uysal S, Baumgärtel V, Lamb DC, Kossiakoff AA, Kotler M, Stroud RM, Marx A, Alian A

Structure 2016 11;24(11):1936-1946

PMID: 27692964

Abstract

HIV-1 integrase (IN) catalyzes viral DNA integration into the host genome and facilitates multifunctional steps including virus particle maturation. Competency of IN to form multimeric assemblies is …

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Specific Recognition of a Single-Stranded RNA Sequence by a Synthetic Antibody Fragment

Shao Y, Huang H, Qin D, Li NS, Koide A, Staley JP, Koide S, Kossiakoff AA, Piccirilli JA

J. Mol. Biol. 2016 10;428(20):4100-4114

PMID: 27593161

Abstract

Antibodies that bind RNA represent an unrealized source of reagents for synthetic biology and for characterizing cellular transcriptomes. However, facile access to RNA-binding antibodies requires the engineering of effective …

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A polar ring endows improved specificity to an antibody fragment

Schaefer ZP, Bailey LJ, Kossiakoff AA

Protein Sci. 2016 07;25(7):1290-8

PMID: 27334407

Abstract

Engineering monovalent Fab fragments into bivalent formats like IgGs or F(ab’)2 can lead to aggregation presumably because of nonspecific off-target interactions that induce aggregation. In an effort to further understand the molecular determinants of nonspecific interactions for engineered antibodies and natively folded …

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Conformational Chaperones for Structural Studies of Membrane Proteins Using Antibody Phage Display with Nanodiscs

Dominik PK, Borowska MT, Dalmas O, Kim SS, Perozo E, Keenan RJ, Kossiakoff AA

Structure 2016 Feb;24(2):300-9

PMID: 26749445

Abstract

A major challenge in membrane biophysics is to define the mechanistic linkages between a protein’s conformational transitions and its function. We describe a novel approach to stabilize transient functional states of membrane proteins in native-like …

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