The phage lambda terminase enzyme: 2. Refolding of the gpNu1 subunit from the detergent-denatured and guanidinium hydrochloride-denatured state yields different oligomerization states and altered protein stabilities

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The terminase enzyme from bacteriophage lambda is responsible for packaging a single genome within the viral capsid. Gold and co-workers have developed a scheme for the solubilization of the small terminase subunit (gpNu1) from inclusion bodies using the strong detergent sarkosyl and purification of the protein to homogeneity (gpNu1 SRK) (Parris et al., J Biol Chem 1994;269:13564–13574). We have developed a similar purification scheme except that guanidinium hydrochloride was used to denature the insoluble protein (gpNu1 GDN). The circular dichroism (CD) spectra of both protein preparations suggest that they are predominantly α-helical when purified and stored in Tris buffers. Moreover, thermal denaturation of the proteins thus purified yielded similar thermodynamic parameters for unfolding ( T m, Δ H m and Δ S m of unfolding of ≈306 K, ≈22 kcal/mol and ≈70 cal/mol·K, respectively). Interestingly, however, when the proteins were purified and stored in imidazole buffers, the gpNu1 SRK preparation lost a significant amount of secondary structure and was more stable to both thermally-induced and guanidinium HCl-induced denaturation than was gpNu1 GDN. The purified gpNu1 monomers oligomerize into apparent tetramers and hexamers in solution and the distribution between these two oligomeric states and into higher order aggregates depends upon buffer composition, salt concentration and protein concentration. Moreover, differences in the oligomerization state of gpNu1 SRK and gpNu1 GDN under identical buffer conditions were observed. The significance of these results with respect to the biological role of the phage lambda gpNu1 protein are discussed.