4.3. Dynamic Light Scattering (DLS)

Aggregation kinetics was registered by an increase in the light-scattering intensity using the Photocor Complex correlation spectrophotometer (PhotoCor Instruments, Inc. College Park, MD, USA) with a He-Ne laser (Model 31-2082, 632.8 nm, 10 mW, Coherent Inc., Santa Clara, CA, USA) as a light source. The scattered light was collected at a 90° angle, and the accumulation time of the autocorrelation functions was 30 s for each experimental point. Glass vials with stoppers were preheated at the desired temperature (50 °C for GDH and 37 °C for Lyz) with the buffer and with/without the crowding agent and/or the osmolyte. Then, the small aliquot of HSPB5 was added, if necessary. In the experiments with Lyz, the protein was also preheated in the vial and the aggregation process was initiated by the addition of DTT to the final concentration of 20 mM. In the experiments with GDH, the aggregation process was initiated by adding the protein into the preheated solution. All experiments were repeated in triplicate.