Differential scanning calorimetry (DSC) has been employed to characterize the thermal denaturation of CPL1 lysozyme and its isolated fragments in the absence and presence of choline. The heat capacity function of CPL1 lysozyme shows two peaks with Tm values of 43.5 and 51.4 degrees C. At saturating concentrations of choline the second transition disappears, and the Tm is shifted to higher temperatures. The DSC thermogram of the C-CPL1 protein corresponding to the carboxyl-terminal domain of CPL1 lysozyme has a single peak with a Tm of 42.9 degrees C. The effect of choline is very similar to that observed for the whole CPL1 lysozyme. The NH2-terminal fragment obtained by proteolytic digestion shows a Tm of 52 degrees C, close to that of 51.4 degrees C found for the second transition of CPL1, and choline does not affect the Tm nor the denaturation enthalpy. These data suggest that choline is bound to the COOH-terminal domain of the protein. Deconvolution of the excess heat capacity curve of the CPL1 lysozyme shows that the data can be fitted to two two-state independent transitions. The analysis of the DSC curves showed that the NH2-terminal unfolding enthalpy steadily decreases with increasing concentrations of choline. These results indicate that, under saturating concentrations of choline, whole CPL1 could unfold as a single cooperative unit. Study holds ProTherm entries: 5144, 5145, 5146 Extra Details: transition 1; C-terminal module; reversibility is from scanning-rate,independent thermogram domain; CPL1 lysozyme; deconvolution;,two-state independent transitions; cooperative unit
Submitter: Connie Wang
Submission Date: April 24, 2018, 8:28 p.m.
|Number of data points||9|
|Proteins||Lysozyme ; Lysozyme|
|Assays/Quantities/Protocols||Experimental Assay: dHcal ; Experimental Assay: Tm ; Experimental Assay: dHvH|
|Libraries||Mutations for sequence VKKNDLFVDVSSHNGYDITGILEQMGTTNTIIKISESTTYLNPCLSAQVEQSNPIGFYHFARFGGDVAEAEREAQFFLDNVPMQVKYLVLDYEDDPSGDAQANTNACLRFMQMIADAGYKPIYYSYKPFTHDNVDYQQILAQFPNSLWIAGYGLNDGTANFEYFPSMDGIRWWQYSSNPFDKNIVLLDDEEDDKPKTAGTWKQDSKGWWFRRNNGSFPYNKWEKIGGVWYYFDSKGYCLTSEWLKDNEKWYYLKDNGAMATGWVLVGSEWYYMDDSGAMVTGWVKYKNNWYYMTNERGNMVSNEFIKSGKGWYFMNTNGELADNPSFTKEPDGLITVA|