Increasing protein stability by altering long-range coulombic interactions.


It is difficult to increase protein stability by adding hydrogen bonds or burying nonpolar surface. The results described here show that reversing the charge on a side chain on the surface of a protein is a useful way of increasing stability. Ribonuclease T1 is an acidic protein with a pI approximately 3.5 and a net charge of approximately -6 at pH 7. The side chain of Asp49 is hyperexposed, not hydrogen bonded, and 8 A from the nearest charged group. The stability of Asp49Ala is 0.5 kcal/mol greater than wild-type at pH 7 and 0.4 kcal/mol less at pH 2.5. The stability of Asp49His is 1.1 kcal/mol greater than wild-type at pH 6, where the histidine 49 side chain (pKa = 7.2) is positively charged. Similar results were obtained with ribonuclease Sa where Asp25Lys is 0.9 kcal/mol and Glu74Lys is 1.1 kcal/mol more stable than the wild-type enzyme. These results suggest that protein stability can be increased by improving the coulombic interactions among charged groups on the protein surface. In addition, the stability of RNase T1 decreases as more hydrophobic aromatic residues are substituted for Ala49, indicating a reverse hydrophobic effect. Study holds ProTherm entries: 6820, 6821, 6822, 6823, 6824, 6825, 6826, 14370, 14371 Extra Details:

Submission Details


Submitter: Connie Wang

Submission Date: April 24, 2018, 8:32 p.m.

Version: 1

Publication Details
Grimsley GR;Shaw KL;Fee LR;Alston RW;Huyghues-Despointes BM;Thurlkill RL;Scholtz JM;Pace CN,Protein Sci. (1999) Increasing protein stability by altering long-range coulombic interactions. PMID:10493585
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 Guanyl-specific ribonuclease Sa P05798 RNSA_KITAU
100.0 Guanyl-specific ribonuclease T1 P00651 RNT1_ASPOR