Browsing by Subject "Self-assembled biomaterials"

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Open Access
Functional bacterial amyloid nanomaterials
(2016-09) Önür, Tuğçe
Mis-folded or unfolded proteins tend to aggregate and aggregated structures are called as amyloids. Amyloid formation contributes to some human diseases and resulting in death in some cases. On the other hand, functional amyloids are found in nature and they are highly ordered assembled structures and they function in cellular events. Some bacteria, fungi or yeast species synthesize these kind of functional amyloids. For example, curli proteins of Escherichia coli play a role in initial attachments for biofilm formation and contribute to stiffness of the biofilm matrix. CsgA is the major subunit while CsgB is the minor subunits which nucleates CsgA polymerization. They are capable of attachment to the abiotic or biotic surfaces. Both of them share some characteristics with non-functional amyloids. For instance, their structures are dominated by ß sheets so they have a rigid amyloid core domain that enables to resist stress factors such as proteases and detergent treatment or pH. Their stable structures and adhesive properties make them useful in materials science. Moreover, high yield could be obtained easily by using molecular biology techniques such as cloning and protein purification so they are highly cost-effective materials. In this study, CsgA and CsgB fibers were proposed as new type of functional biomaterials to do so fiber formations of CsgA and CsgB were analyzed in detail. csgA and csgB genes were cloned into expression vectors. Their ß sheet rich structures were validated with CD analysis and binding capability to Thioflavin T dye were assayed which is the general property of amyloids. Self-seeding and cross-seeding strategies were applied to analyze fiber formation and quartz crystal microbalance with dissipation (QCM-D) was used. Gold coated sensors were deposited with freshly purified proteins and polymerized. Then, sensor surfaces were monitored with SEM and AFM. With self-seeding strategies long and branched fibers were obtained from CsgA proteins while sphere like structures were formed by CsgB proteins. Also, it was concluded from the cross-seeding experiments, the order of protein addition determines the final assembled structures. Furthermore, fluorescent properties of CsgA and CsgB were analyzed in detail for the first time. Finally, binding affinity of the purified proteins to different materials (gold, silica and hydroxyapatite) were determined by using QCM-D.