The Influence of the Mixed DPC:SDS Micelle on the Structure and Oligomerization Process of the Human Cystatin C

Abstract

Human cystatin C (hCC), a member of the superfamily of papain-like cysteine protease inhibitors, is the most widespread cystatin in human body fluids. Physiologically active hCC is a monomer, which dimerization and oligomerization lead to the formation of the inactive, insoluble amyloid form of the protein, strictly associated with cerebral amyloid angiopathy, a severe state causing death among young patients. It is known, that biological membranes may accelerate the oligomerization processes of amyloidogenic proteins. Therefore, in this study, we describe an influence of membrane mimetic environment—mixed dodecylphosphocholine:sodium dodecyl sulfate (DPC:SDS) micelle (molar ratio 5:1)—on the effect of the hCC oligomerization. The hCC–micelle interactions were analyzed with size exclusion chromatography, circular dichroism, and nuclear magnetic resonance spectroscopy. The experiments were performed on the wild-type (WT) cystatin C, and two hCC variants—V57P and V57G. Collected experimental data were supplemented with molecular dynamic simulations, making it possible to highlight the binding interface and select the residues involved in interactions with the micelle. Obtained data shows that the mixed DPC:SDS micelle does not accelerate the oligomerization of protein and even reverses the hCC dimerization process.

Copper interaction with cystatin C: effects on protein structure and oligomerization

“…Therefore, a more stable V57G hCC variant was used in these experiments [11,39,40]. Its 2D 1 H‐ 15 N HSQC spectrum is identical to that obtained previously [11,39,40]. The addition of CuCl 2 resulted in broadened or disappeared signals from several residues, but the overall cross‐peak pattern was not perturbed (Fig.…”

“…Recording the NMR spectra of the wild‐type hCC is troublesome due to concomitant protein dimerization. Therefore, a more stable V57G hCC variant was used in these experiments [11,39,40]. Its 2D 1 H‐ 15 N HSQC spectrum is identical to that obtained previously [11,39,40].…”

Binding of Different Cyclosporin Variants to Micelles Evidenced by NMR and MD Simulations

“…Perdeuterated dodecylphosphocholine was specifically developed as a membrane-like detergent for use in NMR experiments. The predominant phospholipid in animal (but not bacterial) cell membranes, DPC is nevertheless a true detergent with properties very similar to SDS [ 16 ]. To date, DPC is one of the most thoroughly studied model systems for studying lipid-binding peptides and proteins.…”