title: Contributions of folding cores to the thermostabilities of two ribonucleases H.
abstract: To investigate the contribution of the folding cores to the thermodynamic stability of RNases H, we used rational design to create two chimeras composed of parts of a thermophilic and a mesophilic RNase H. Each chimera combines the folding core from one parent protein and the remaining parts of the other. Both chimeras form active, well-folded RNases H. Stability curves, based on CD-monitored chemical denaturations, show that the chimera with the thermophilic core is more stable, has a higher midpoint of thermal denaturation, and a lower change in heat capacity (DeltaCp) upon unfolding than the chimera with the mesophilic core. A possible explanation for the low DeltaCp of both the parent thermophilic RNase H and the chimera with the thermophilic core is the residual structure in the denatured state. On the basis of the studied parameters, the chimera with the thermophilic core resembles a true thermophilic protein. Our results suggest that the folding core plays an essential role in conferring thermodynamic parameters to RNases H.
authors.affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, California 94720, USA.
authors.forename: Srebrenka
authors.initials: S
authors.lastname: Robic
authors.forename: James M
authors.initials: JM
authors.lastname: Berger
authors.forename: Susan
authors.initials: S
authors.lastname: Marqusee
mesh_terms: D000595:Amino Acid Sequence; D002678:Chimera; D002942:Circular Dichroism; D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology; D006358:Hot Temperature; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010957:Plasmids; D016133:Polymerase Chain Reaction; D011489:Protein Denaturation; D017510:Protein Folding; D016914:Ribonuclease H / Q000737:chemistry*; D017386:Sequence Homology, Amino Acid; D013816:Thermodynamics; D016963:Thermus thermophilus / Q000201:enzymology*; D014961:X-Ray Diffraction
chemical_list: D016914:Ribonuclease H