1.

The mechanism of amyloid fibril formation by stefin B: temperature and protein concentration dependence of the rates.

Cystatins have proved to be useful model system to study amyloidogenesis. We have extended previous studies of the kinetics of amyloid-fibril formation by human stefin B and proposed an improved model for the reaction. The observed kinetics follow the nucleation and growth behavior observed for many other amyloidogenic proteins. The minimal kinetic scheme includes nucleation, fibril growth and nonproductive oligomerization, best explained by an off-pathway state with a rate-limiting escape rate.

COBISS.SI-ID: 21880103

2.

Monitoring compartment-specific substarte cleavage by cathepsins B, K, L and S at physiological pH and redox conditions.

Cysteine cathepsins are known to primarily cleave their substrates at reducing and acidic conditions within endo-lysosomes. They have also been linked to extracellular proteolysis. We developed an assay to test for proteolytic processing of a natural substrate by cysteine cathepsins in the extracellular space. Our study emphasizes that the proteolytic functions of cysteine cathepsins in the thyroid are not restricted to endo-lysosomes but include pivotal roles in extracellular substrate utilization. We have approached this by simulating physiological conditions in protease activity assays.

COBISS.SI-ID: 23357479

3.

Averaged kick maps: less noise, more signal... and probably less bias.

Use of reliable density maps is crucial for rapid and successful crystal structure determination. Here, the averaged kick (AK) map approach is investigated, its application is generalized and it is compared with other map-calculation methods. The conclusion is that AK maps can be useful throughout the entire progress of crystal structure determination, offering the possibility of improved map interpretation.

COBISS.SI-ID: 22793511

4.

Essential role of Pro 74 in stefin B amyloid fibril formation: dual action of cyclophilin A on the process.

The stefin B P74S mutant did not fibrillate over the time of observation at 25 C, and it exhibited a prolonged lag phase at 30 C and 37 C. The peptidyl prolyl cis/trans isomerase cyclophilin A prolonged the lag phase and increased the yield and length of the fibrils. Addition of the inactive cyclophilin A R55A variant still resulted in a prolonged lag phase but did not mediate the increase of the final fibril yield. These results demonstrate that peptidyl prolyl cis/trans isomerism is rate-limiting in stefin B fibril formation.

COBISS.SI-ID: 22481191

5.

Autocatalytic processing of procathepsin B is triggered by proenzyme activity.

Cathepsin B and other cysteine proteases are synthesized as zymogens, which are processed to their mature forms autocatalytically or by other proteases. Initiation of the autocatalytic processing has not yet been clarified. A model for autocatalytic activation of cysteine cathepsins is proposed, involving propeptide dissociation from the active-site cleft, followed by a bimolecular proteolytic removal of the propeptide. Such activation, may have important physiological consequences because cathepsin zymogens were often found secreted in various pathological states.

COBISS.SI-ID: 22392615