• Derick Shu Fuh
4. semester, Nanobioteknologi (cand.polyt.), Kandidat (Kandidatuddannelse)
One of the key challenges in nanoscience is the technological utilization of self-assembly systems; a process wherein molecules will spontaneously associate under equilibrium conditions into reproducible aggregates of supramolecules. Supramolecular structures and devices in the dimensions of a few to tens of nanometers have been developed using crystalline bacterial cell surface proteins (S-layer proteins). Treatment of whole bacterial cells with detergents and chaotropic agents enable the isolation of S-layer proteins of different molecular weights, having the intrinsic property to recrystallize into two dimensional arrays at a broad spectrum of surfaces and interfaces. The well-defined arrangement of functional groups of the S-layers on solid support permits the binding of molecules including enzymes in defined regular arrays. This can be applied in the nanofabrication of analytical based biosensors among other applications. The immobilization of enzymes facilitates the separation of the enzyme from the reaction mixture and can significantly lower the cost of the enzyme.
The aim of this project is to isolate and purify crystalline surface layer proteins from the bacteria cell envelope of a Gram positive bacteria; Corynebacterium glutamicum using a detergent (SDS) and high molar concentrations of a chaotropic agent (guanidine hydrochloride), characterize the isolated S-layer proteins by SDS PAGE in order to determine its molecular weight and do an AFM analysis of the self-assembled S-layers on a mica surface and finally immobilize a hydrolytic enzyme; cutinase on the self-assembled S-layer proteins. An esterase activity test for cutinase will be used to study the binding of cutinase to the self-assembled S-layers on the mica surface. The immobilized cutinase activity will be compared with the activity of the enzyme in solution.
Key Words: Self-assembly, Surface Layer Proteins, Enzyme Immobilization, Cutinase, Atomic Force Microscopy
Udgivelsesdato2 jun. 2014
Antal sider58
ID: 198671515