Biophysical studies on the full-length human cyclin A(2): Protein stability and folding/unfolding thermodynamics

Human cyclin A(2) participates in cell cycle regulation, DNA replication, and transcription. Its overexpression has been implicated in the development and progression of a variety of human cancers. However, cyclin A(2) or its truncated form is very unstable in the absence of binding partner, which makes it difficult to get a deep insight of structural basis of the interactions. Therefore, biophysical studies of the full-length human cyclin A, would provide important information regarding protein stability and folding/unfolding process.

Evaluation of different culture conditions for high-level soluble expression of human cyclin A(2) with pET vector in BL21 (DE3) and spectroscopic characterization of its inclusion body structure

In this paper, we evaluated various parameters of culture condition affecting high-level soluble expression of human cyclin A, in Escherichia coli BL21(DE3), and demonstrated that the highest protein yield was obtained using TB(no glycerol) + 0.5% glucose medium at 25 degrees C. By single immobilized metal ion affinity chromatography, we got highly purified human cyclin A(2) with a yield ranged from 20 to 30 mg/L. By amyloid-diagnostic dye ThT binding and Fourier transform infrared spectroscopy, we observed a significant decrease in alpha-helix content and an increase in beta-sheet structure in cyclin A(2) inclusion body in comparison to its native protein, and confirmed the resemblance of the internal organization of cyclin A(2) inclusion body and amyloid fibrils.