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Biomolecules 2017, 7(3), 57; doi:10.3390/biom7030057

Metal Binding Properties of the N-Terminus of the Functional Amyloid Orb2

1
Department of Biochemistry & Molecular Medicine and the Zilkha Neurogenetic Institute, Keck School of Medicine of USC, Los Angeles, CA 90033, USA
2
Department of Biomedical Engineering, McGill University, Montreal, QC H3A 2B4, Canada
3
U.S. Army Construction Engineering Research Laboratory, 2902 Newmark Dr, Champaign, IL 61822, USA
*
Author to whom correspondence should be addressed.
Academic Editors: Margaret Sunde, Matthew Chapman and Sarah Perrett
Received: 3 June 2017 / Revised: 14 July 2017 / Accepted: 21 July 2017 / Published: 1 August 2017
(This article belongs to the Special Issue Functional Amyloids)
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Abstract

The cytoplasmic polyadenylation element binding protein (CPEB) homologue Orb2 is a functional amyloid that plays a key regulatory role for long-term memory in Drosophila. Orb2 has a glutamine, histidine-rich (Q/H-rich) domain that resembles the Q/H-rich, metal binding domain of the Hpn-like protein (Hpnl) found in Helicobacter pylori. In the present study, we used chromatography and isothermal titration calorimetry (ITC) to show that the Q/H-rich domain of Orb2 binds Ni2+ and other transition metals ions with μM affinity. Using site directed mutagenesis, we show that several histidine residues are important for binding. In particular, the H61Y mutation, which was previously shown to affect the aggregation of Orb2 in cell culture, completely inhibited metal binding of Orb2. Finally, we used thioflavin T fluorescence and electron microscopy images to show that Ni2+ binding induces the aggregating of Orb2 into structures that are distinct from the amyloid fibrils formed in the absence of Ni2+. These data suggest that transition metal binding might be important for the function of Orb2 and potentially long-term memory in Drosophila. View Full-Text
Keywords: amyloid; protein–metal interaction; aggregation; ITC; thioflavin T fluorescence amyloid; protein–metal interaction; aggregation; ITC; thioflavin T fluorescence
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MDPI and ACS Style

Bajakian, T.H.; Cervantes, S.A.; Soria, M.A.; Beaugrand, M.; Kim, J.Y.; Service, R.J.; Siemer, A.B. Metal Binding Properties of the N-Terminus of the Functional Amyloid Orb2. Biomolecules 2017, 7, 57.

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