Mutations in the cystatin B gene cause a rare form of epilepsy EPM1. Its two missense mutants, G50E and Q71P, lack the inhibitory activity and are partially unfolded, which leads to changes in their aggregation behavior, both in vitro and in the cell. SDS-PAGE and MALDI-TOF mass spectrometry were used to follow the hydrolysis of human stefin B wild type, G50E and Q71P by cathepsins B and S in vitro. Cathepsin S was found to degrade both mutants with Q71P being degraded faster. This correlates with the openness of the protein structure, Q71P having more exposed hydrophobic surfaces. Cathepsin B acted more selectively, degrading G50E into smaller fragments, while Q71P was cleaved only at the exposed N-terminal end.
COBISS.SI-ID: 26511655
Unlike a number of amyloid-forming proteins, stefin B (cystatin B) forms amyloids under conditions where the native state predominates. In order to trigger oligomerization processes, the stability of the protein needs to be compromised, favoring structural re-arrangement, however, accelerating fibril formation is not a simple function of protein stability. We report here on how optimal conditions for amyloid formation lead to the destabilization of dimeric and tetrameric states of the protein in favor of the monomer. Small, highly localized structural changes can be mapped out that allow us to visualize directly areas of the protein which eventually become responsible for triggering amyloid formation. These regions of the protein overlap with the Cu (II)-binding sites which we futher identify here. Stefin B was previously reported as a copper-binding protein (Zerovnik et al., 2006).
COBISS.SI-ID: 26390823
It is well documented that amyloid forming peptides and proteins interact with membranes and that this correlates with cytotoxicity. To introduce the theme we give a brief description of some amyloidogenic proteins and note their similarities with pore forming toxins (PFTs) and cell penetrating peptides. Human stefin B, a member of the family of cystatins, is an amyloidogenic protein in vitro. This review describes our studies of the interaction of stefin B oligomers and prefibrillar aggregates with model membranes leading to pore formation. We discuss the possible functional and pathological significance of such pores formed by human stefin B.
COBISS.SI-ID: 5029658