Stability and cytotoxicity of crystallin amyloid nanofibrils

Abstract

Previous work has identified crystallin proteins extracted from fish eye lenses as a cheap and readily available source for the self-assembly of amyloid nanofibrils. However, before exploring potential applications, the biophysical aspects and safety of this bionanomaterial need to be assessed so as to ensure that it can be effectively and safely used. In this study, crude crystallin amyloid fibrils are shown to be stable across a wide pH range, in a number of industrially relevant solvents, at both low and high temperatures, and in the presence of proteases. Crystallin nanofibrils were compared to well characterised insulin and whey protein fibrils using Thioflavin T assays and TEM imaging. Cell cytotoxicity assays suggest no adverse impact of both mature and fragmented crystallin fibrils on cell viability of Hec-1a endometrial cells. An IR microspectroscopy study supports long-term structural integrity of crystallin nanofibrils.

Amyloid Fibrils from Hemoglobin

“…These results were in agreement with the previous studies, which have demonstrated the stability of whey, insulin, and crystallin fibrils in MeOH, EtOH, and MeCH(OH)Me [10,23]. As previously observed, the decrease in ThT fluorescence at 24 h could be linked to the degradation or solubilisation of intact AHB fibrils [41].…”

“…AHB fibrils were not stable in DMSO, in agreement with the literature on fibrils derived from other protein sources [23]. The dissolution of AHB fibrils by DMSO is consistent with the disruption of the hydrogen bond network, which stabilizes the fibril structure [42].…”

“…In this study, we investigated AHB fibril stability at −20 °C, 4 °C, 22 °C (room temperature), 37 °C, and 80 °C, following temperatures investigated in the literature by Kaur and co-workers [23]. AHB fibrils were stable at all temperatures, as suggested by the maintenance of ThT fluorescence (Figure 5A).…”

“…Previous research has demonstrated the functionalization of fibrils for various applications such as biosensors [9,10,11,12,13], nanowires [14,15,16], nanocomposites [17,18], thin films [19], nanoporous matrices [20], hydrogels [21], and aerogels [22]. Their stability over a broad-range of temperatures, pHs, solvents, and proteases, allows amyloid fibrils to be exploited for many applications [13,23]. …”

Evaluation of protease resistance and toxicity of amyloid-like food fibrils from whey, soy, kidney bean, and egg white

“…The Hec-1a cell line is an immortal epithelial cell line derived from endometrial tissue (Kamat et al, 2007). In a parallel study assessing the use of fibrils as nanomaterials, we recently investigated the toxicity of WPI fibrils towards the endothelial cell line Hec-1a and confirmed that WPI fibrils were non-toxic to this cell line (Kaur et al, 2014). To our knowledge, the potential toxicity of WPI, KPI, SPI, and OVA fibrils has not been addressed previously in the context of food.…”