Contribution of conformational stability and reversibility of unfolding to the increased thermostability of human and bovine superoxide dismutase mutated at free cysteines.


The conformational stability and reversibility of unfolding of the human dimeric enzyme Cu Zn superoxide dismutase (HSOD) and the three mutant enzymes constructed by replacement of Cys6 by Ala and Cys111 by Ser, singly and in combination, were determined by differential scanning calorimetry. The differential scanning calorimetry profile of wild-type HSOD consists of two components, which probably represent the unfolding of the oxidized and reduced forms of the enzyme, with denaturation temperatures (Tm) of 74.9 and 83.6 degrees C, approximately 7 degrees lower than those for bovine superoxide dismutase (BSOD). The conformational stabilities of the two components of the mutant HSOD's differ only slightly from those of the wild type (delta delta Gs of -0.2 to +0.8 kcal/mol of dimer), while replacement of the BSOD Cys6 by Ala is somewhat destabilizing (delta delta G of -0.7 to -1.3 kcal/mol of dimer). These small alterations in conformational stability do not correlate with the large increases in resistance to thermal inactivation following substitution of free Cys in both HSOD and BSOD (McRee, D.E., Redford, S.M., Getzoff, E.D., Lepock, J.R., Hallewell, R.A., and Tainer, J.A. (1990) J. Biol. Chem. 265, 14234-14241 and Hallewell, R.A., Imlay, K.C., Laria, I., Gallegos, C., Fong, N., Irvine, B., Getzoff, E.D., Tainer, J.A., Cubelli, D.E., Bielski, B.H.J., Olson, P., Mallenbach, G.T., and Cousens, L.S. (1991) Proteins Struct. Funct. Genet., submitted for publication). The reversibility of unfolding was determined by scanning part way through the profile, cooling, rescanning, and calculating the amount of protein irreversibly unfolded by the first scan. The order of reversibility at a constant level of unfolding is the same as the order of resistance to inactivation for both the HSOD and BSOD wild-type and mutant enzymes. Thus, the greater resistance to thermal inactivation of the superoxide dismutase enzymes with free Cys replaced by Ala or Ser is dominated by a greater resistance to irreversible unfolding and relatively unaffected by changes in conformational stability. Study holds ProTherm entries: 5179, 5180, 5181, 5182, 5183 Extra Details: free cysteines; conformational stability; reversibility;,thermal inactivation

Submission Details

ID: yq6BcGwJ3

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:29 p.m.

Version: 1

Publication Details
Lepock JR;Frey HE;Hallewell RA,J. Biol. Chem. (1990) Contribution of conformational stability and reversibility of unfolding to the increased thermostability of human and bovine superoxide dismutase mutated at free cysteines. PMID:2254318
Additional Information

Structure view and single mutant data analysis

Study data

No weblogo for data of varying length.
Colors: D E R H K S T N Q A V I L M F Y W C G P

Data Distribution

Studies with similar sequences (approximate matches)

Correlation with other assays (exact sequence matches)

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
100.0 Superoxide dismutase [Cu-Zn] P00442 SODC_BOVIN
98.7 Superoxide dismutase [Cu-Zn] Q52RN5 SODC_BOSMU
98.0 Superoxide dismutase [Cu-Zn] Q5FB29 SODC_CAPHI
97.4 Superoxide dismutase [Cu-Zn] P09670 SODC_SHEEP
94.7 Superoxide dismutase [Cu-Zn] O46412 SODC_CEREL
100.0 Superoxide dismutase [Cu-Zn] P00441 SODC_HUMAN
100.0 Superoxide dismutase [Cu-Zn] P60052 SODC_PANTR
95.5 Superoxide dismutase [Cu-Zn] Q8HXQ4 SODC_PONPY
94.2 Superoxide dismutase [Cu-Zn] Q8HXQ3 SODC_HYLLA
93.5 Superoxide dismutase [Cu-Zn] Q8HXP9 SODC_SAPAP
92.2 Superoxide dismutase [Cu-Zn] Q8HXP8 SODC_CALJA
91.6 Superoxide dismutase [Cu-Zn] Q8HXQ1 SODC_MACFA
91.6 Superoxide dismutase [Cu-Zn] Q8HXQ2 SODC_MACFU
91.6 Superoxide dismutase [Cu-Zn] Q8HXQ0 SODC_MACMU