Generation and characterization of novel conformation-specific monoclonal antibodies for α-synuclein pathology

Vaikath, Nishant N.; Majbour, Nour K.; Paleologou, Katerina E.; Ardah, Mustafa T.; Dam, Esther van; Berg, Wilma D. J. van de; Forrest, Shelley L.; Parkkinen, Laura; Gai, Wei-Ping; Hattori, Nobutaka; Takanashi, Masashi; Lee, Seung-Jae; Mann, David; Imai, Yuzuru; Halliday, Glenda M.; Li, Jiayi; El-Agnaf, Omar M. A.

doi:10.1016/j.nbd.2015.04.009

Cited by 125 publications

(150 citation statements)

References 66 publications

Supporting

Mentioning

136

Contrasting

Unclassified

Order By: Relevance

“…Shannon and colleagues reporting an incidence of 8% used an antibody directed to the C-terminus of αS [14]. The significance of the antibody used was also recently emphasized in the work by Vaikath and colleagues [16]. It is worth noting that most studies reporting low incidence (0-8%) of αS-IR in the GI tract, in line with the work of Shannon and colleagues, have been carried out on superficial endoscopic biopsies lacking myenteric plexus [13,14].…”

Section: Discussionmentioning

confidence: 89%

Neuronal Protein Alteration in Enteric Dysmotility Syndrome

Alafuzoff¹,

Popova²

2016

J Alzheimers Dis Parkinsonism

View full text Add to dashboard Cite

Little is known about the enteric ganglionic system in subjects with gastrointestinal dysmotility syndrome (GIDS). Furthermore, dysfunction of gastrointestinal motility is an early complaint of subjects with Parkinson's disease.Here, we assessed p62/sequestosome-1(p62) and α-synuclein (αS) immunoreactivity (IR) in full-thickness bowel specimens of the gut obtained from six subjects with GIDS and from 17 controls.

show abstract

Section: Discussionmentioning

confidence: 89%

Neuronal Protein Alteration in Enteric Dysmotility Syndrome

Alafuzoff¹,

Popova²

2016

J Alzheimers Dis Parkinsonism

View full text Add to dashboard Cite

show abstract

“…Given these challenges, antibodies with conformational specificity for prefibrillar oligomers and amyloid fibrils have proven valuable for biochemical characterization (17)(18)(19)(20)(21)(22)(23)(24)(25)(26)(27)(28). An obvious strength of such antibodies is their ability to detect specific types of aggregates formed both in vivo and in vitro.…”

mentioning

confidence: 99%

“…The most widely used one is immunization, which has yielded a wide range of important antibodies specific for different types of prefibrillar oligomers and amyloid fibrils (17)(18)(19)(20)(25)(26)(27). Another powerful approach that has proven useful is to employ in vitro display methods such as phage and yeast surface display (21-24, 29 -37).…”

mentioning

confidence: 99%

Design and Optimization of Anti-amyloid Domain Antibodies Specific for β-Amyloid and Islet Amyloid Polypeptide

Lee

Julian

Tiller

et al. 2016

Journal of Biological Chemistry

View full text Add to dashboard Cite

Antibodies with conformational specificity are important for detecting and interfering with polypeptide aggregation linked to several human disorders. We are developing a motif-grafting approach for designing lead antibody candidates specific for amyloid-forming polypeptides such as the Alzheimer peptide (A␤). This approach involves grafting amyloidogenic peptide segments into the complementarity-determining regions (CDRs) of single-domain (V H ) antibodies. Here we have investigated the impact of polar mutations inserted at the edges of a large hydrophobic A␤42 peptide segment (A␤ residues 17-42) in CDR3 on the solubility and conformational specificity of the corresponding V H domains. We find that V H expression and solubility are strongly enhanced by introducing multiple negatively charged or asparagine residues at the edges of CDR3, whereas other polar mutations are less effective (glutamine and serine) or ineffective (threonine, lysine, and arginine). Moreover, A␤ V H domains with negatively charged CDR3 mutations show significant preference for recognizing A␤ fibrils relative to A␤ monomers, whereas the same V H domains with other polar CDR3 mutations recognize both A␤ conformers. We observe similar behavior for a V H domain grafted with a large hydrophobic peptide from islet amyloid polypeptide (residues 8 -37) that contains negatively charged mutations at the edges of CDR3. These findings highlight the sensitivity of antibody binding and solubility to residues at the edges of CDRs, and provide guidelines for designing other grafted antibody fragments with hydrophobic binding loops.The misfolding and assembly of peptides and proteins into prefibrillar oligomers and amyloid fibrils is linked to several neurodegenerative diseases (1-4). To understand the contributions of polypeptide aggregation to such disorders, it is important to characterize the biochemical properties of aggregates. However, pre-amyloid and amyloid aggregates are difficult to characterize using many traditional biochemical methods that are routinely used for soluble proteins. High-resolution structural analysis of such aggregates is particularly challenging and must be performed using specialized methods such as solid state NMR (5-13) or x-ray crystallography of small peptide fragments (14 -16). These powerful structural methods generally lack the time resolution to probe pre-amyloid intermediates and oligomers unless they can be kinetically trapped.Given these challenges, antibodies with conformational specificity for prefibrillar oligomers and amyloid fibrils have proven valuable for biochemical characterization (17-28). An obvious strength of such antibodies is their ability to detect specific types of aggregates formed both in vivo and in vitro. The ability of conformational antibodies to bridge in vivo and in vitro studies is important for understanding the biochemical mechanisms that contribute to protein misfolding disorders.Several approaches have been used to generate conformational antibodies. The most widely used one is immuni...

show abstract

“…The Syn-O1 antibody was a monoclonal raised in mice against α-synuclein fibrils (30). The antibody selectively binds α-synuclein oligomers and fibrils and not α-synuclein monomers on inhibition ELISA and dot blotting (30).…”

Section: Syn-o1mentioning

confidence: 99%

“…We used a conventional antibody against α-synuclein in addition to novel conformation-specific antibodies against α-synuclein oligomers and fibrils (30).…”

Section: Introductionmentioning

confidence: 99%

Rotenone and elevated extracellular potassium concentration induce cell‐specific fibrillation of α‐synuclein in axons of cholinergic enteric neurons in the guinea‐pig ileum

Sharrad

Chen

Gai

et al. 2016

Neurogastroenterology Motil

View full text Add to dashboard Cite

Background Parkinson's disease is a progressive neurodegenerative disorder that results in the widespread loss of select classes of neurons throughout the nervous system. The pathological hallmarks of Parkinson's disease are Lewy bodies and neurites, of which α‐synuclein fibrils are the major component. α‐Synuclein aggregation has been reported in the gut of Parkinson's disease patients, even up to a decade before motor symptoms, and similar observations have been made in animal models of disease. However, unlike the central nervous system, the nature of α‐synuclein species that form these aggregates and the classes of neurons affected in the gut are unclear. We have previously reported selective expression of α‐synuclein in cholinergic neurons in the gut (J Comp Neurol. 2013; 521:657), suggesting they may be particularly vulnerable to degeneration in Parkinson's disease. Methods In this study, we used immunohistochemistry to detect α‐synuclein oligomers and fibrils via conformation‐specific antibodies after rotenone treatment or prolonged exposure to high [K+] in ex vivo segments of guinea‐pig ileum maintained in organotypic culture. Key Results Rotenone and prolonged raising of [K+] caused accumulation of α‐synuclein fibrils in the axons of cholinergic enteric neurons. This took place in a time‐ and, in the case of rotenone, concentration‐dependent manner. Rotenone also caused selective necrosis, indicated by increased cellular autofluorescence, of cholinergic enteric neurons, labeled by ChAT‐immunoreactivity, also in a concentration‐dependent manner. Conclusions & Inferences To our knowledge, this is the first report of rotenone causing selective loss of a neurochemical class in the enteric nervous system. Cholinergic enteric neurons may be particularly susceptible to Lewy pathology and degeneration in Parkinson's disease.

show abstract

Generation and characterization of novel conformation-specific monoclonal antibodies for α-synuclein pathology

Cited by 125 publications

References 66 publications

Neuronal Protein Alteration in Enteric Dysmotility Syndrome

Neuronal Protein Alteration in Enteric Dysmotility Syndrome

Design and Optimization of Anti-amyloid Domain Antibodies Specific for β-Amyloid and Islet Amyloid Polypeptide

Rotenone and elevated extracellular potassium concentration induce cell‐specific fibrillation of α‐synuclein in axons of cholinergic enteric neurons in the guinea‐pig ileum

Contact Info

Product

Resources

About