Polyamines putrescine and spermidine as modulators of protein aggregation rate: The effect on DTT-induced aggregation of α-lactalbumin.

Protein aggregation is undesirable for cells due to its possible toxicity, and is also undesirable in biotechnology and pharmaceuticals. Polyamines are known to be capable of both suppressing and stimulating protein aggregation. In the present work polyamines (spermidine, putrescine) have been shown to alter the pathway of α-lactalbumin aggregation induced by dithiothreitol, leading to the formation of larger protein particles during the initial stages of aggregation and promoting the later stage of sticking of aggregates. According to the aggregation kinetics data, polyamines accelerate protein aggregation in a concentration-dependent manner, with a maximum at 50 mM spermidine and 100 mM putrescine. With a further increase in polyamines concentration the effect of aggregation acceleration decreased, thus, the modulation of the aggregation rate by polyamines was shown. A comparison of the aggregation kinetics and hydrodynamic radii growth data registered by dynamic light scattering with the data obtained by asymmetric flow field-flow fractionation and analytical ultracentrifugation allowed us to describe the early stages of aggregation and formation of initial α-lactalbumin clusters. Our results provide a deeper insight into the mechanism of amorphous aggregation of α-lactalbumin and polyamines action on protein aggregation and protein-protein interaction in general.

Functional and Structural Properties of Cytoplasmic Tropomyosin Isoforms Tpm1.8 and Tpm1.9.

The actin cytoskeleton is one of the most important players in cell motility, adhesion, division, and functioning. The regulation of specific microfilament formation largely determines cellular functions. The main actin-binding protein in animal cells is tropomyosin (Tpm). The unique structural and functional diversity of microfilaments is achieved through the diversity of Tpm isoforms. In our work, we studied the properties of the cytoplasmic isoforms Tpm1.8 and Tpm1.9. The results showed that these isoforms are highly thermostable and differ in the stability of their central and C-terminal fragments. The properties of these isoforms were largely determined by the 6th exons. Thus, the strength of the end-to-end interactions, as well as the affinity of the Tpm molecule for F-actin, differed between the Tpm1.8 and Tpm1.9 isoforms. They were determined by whether an alternative internal exon, 6a or 6b, was included in the Tpm isoform structure. The strong interactions of the Tpm1.8 and Tpm1.9 isoforms with F-actin led to the formation of rigid actin filaments, the stiffness of which was measured using an optical trap. It is quite possible that the structural and functional features of the Tpm isoforms largely determine the appearance of these isoforms in the rigid actin structures of the cell cortex.