Insulin fibrillation: The influence and coordination of Zn 2+

Abstract

Protein amyloid fibrillation is obtaining much focus because it is connected with amyloid-related human diseases such as Alzheimer's disease, diabetes mellitus type 2, or Parkinson's disease. The influence of metal ions on the fibrillation process and whether it is implemented in the amyloid fibrils has been debated for some years. We have therefore investigated the influence and binding geometry of zinc in fibrillated insulin using extended X-ray absorption fine-structure and X-ray absorption near-edge structure spectroscopy. The results were validated with fibre diffraction, Transmission Electron Microscopy and Thioflavin T fluorescence measurements. It is well-known that Zn ions coordinate and stabilize the hexameric forms of insulin. However, this study is the first to show that zinc indeed binds to the insulin fibrils. Furthermore, zinc influences the kinetics and the morphology of the fibrils. It also shows that zinc coordinates to histidine residues in an environment, which is similar to the coordination seen in the insulin R hexamers, where three histidine residues and a chloride ion is coordinating the zinc.

Solution structures of long-acting insulin analogues and their complexes with albumin

“…The different multihexamerizations indicate that their modes of hexamer-hexamer association are fundamentally different. The driving force of association results from their fatty-acid moieties, as human insulin is normally observed in an equilibrium between monomer, dimer and hexamer (Frankaer et al, 2017;Jorgensen et al, 2011). While detemir has a C14 fatty acid attached to LysB29, the second-generation product degludec has a C16 dicarboxylic fatty acid attached through a -glutamate linker.…”

Prion-derived tetrapeptide stabilizes thermolabile insulin via conformational trapping

“…Availability of injectable insulin formulation has been a breakthrough in diabetes management in achieving long-term glycemic control and preventing complications ( Baram et al., 2018 ; Heller et al, 2007 ; Moroder and Musiol, 2017 ; Owens et al., 2001 ; Xiong et al., 2019 ; Zaykov et al., 2016 ); it still, however, suffers from certain disadvantages including temperature-sensitive fibrillation in solution and development of subcutaneous tumor-like mass designated as “amyloidoma” at the site of injection ( Hua and Weiss, 2004 ; Ivanova et al., 2009 ; Nilsson, 2016 ; Woods et al., 2012 ; Yumlu et al, 2009 ). Worldwide efforts were thus made to develop thermostable insulin either by making recombinant insulin species with mutations or stabilizing native insulin with salts, Zn 2+ ions, and small molecules such as meta-cresol ( Frankær et al., 2017 ; Gong et al., 2014 ; Han et al., 2017 ; Kachooei et al, 2014 ; Lee et al, 2014 ; Patel et al, 2018 ; Saithong et al, 2018 ; Wang et al, 2011 ; Zheng and Lazo, 2018 ). However, small molecules are found to be toxic in long-term usage and are inefficient in optimally preventing fibrillation ( Teska et al., 2014 ; Weber et al., 2015 ).…”

FT-IR versus EC-QCL spectroscopy for biopharmaceutical quality assessment with focus on insulin—total protein assay and secondary structure analysis using attenuated total reflection

“…These adverse storage conditions can be associated with breaking the storage cold chain or during transport, encountered with shaking and elevated temperatures [1]. First deteriorating effects can be protein misfolding with drastic changes in the protein secondary structure, followed by protein aggregation and fibril forming leading to a reduction in the potency of the active pharmaceutical ingredient [2][3][4]. Aggregation can be followed by size-exclusion chromatography (SEC) [5] or by sodium dodecyl polyacrylamide gel electrophoresis (SDS-PAGE) [6].…”

“…Early events in the globular insulin peptide's misfolding during elevated stress and temperature events reveal changes in the peptide's A and B chains, including the exposure of partially unfolded sites leading to intermediate monomeric states. These intermediates are enabling subsequent formation of beta sheet substructures and can be detected without changes concerning their amino acid sequence [3,4]. Due to this possibility, for example, HPLC methods are able to detect both types of active and inactive insulins, but without discriminating between the two.…”