Contribution of histidine residues to the conformational stability of ribonuclease T1 and mutant Glu-58----Ala.


Abstract

The pK values of the histidine residues in ribonuclease T1 (RNase T1) are unusually high: 7.8 (His-92), 7.9 (His-40), and 7.3 (His-27) [Inagaki et al. (1981) J. Biochem. 89, 1185-1195]. In the RNase T1 mutant Glu-58----Ala, the first two pK values are reduced to 7.4 (His-92) and 7.1 (His-40). These lower pKs were expected since His-92 (5.5 A) and His-40 (3.7 A) are in close proximity to Glu-58 at the active site. The conformational stability of RNase T1 increases by over 4 kcal/mol between pH 9 and 5, and this can be entirely accounted for by the greater affinity for protons by the His residues in the folded protein (average pK = 7.6) than in the unfolded protein (pk approximately 6.6). Thus, almost half of the net conformational stability of RNase T1 results from a difference between the pK values of the histidine residues in the folded and unfolded conformations. In the Glu-58----Ala mutant, the increase in stability between pH 9 and 5 is halved (approximately 2 kcal/mol), as expected on the basis of the lower pK values for the His residues in the folded protein (average pK = 7.1). As a consequence, RNase T1 is more stable than the mutant below pH 7.5, and less stable above pH 7.5. These results emphasize the importance of measuring the conformational stability as a function of pH when comparing proteins differing in structure. Study holds ProTherm entries: 2569, 2570, 2571, 2572 Extra Details: Ribonuclease T1; conformational stability; histidine; affinity

Submission Details

ID: Nft3tjcd3

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:19 p.m.

Version: 1

Publication Details
McNutt M;Mullins LS;Raushel FM;Pace CN,Biochemistry (1990) Contribution of histidine residues to the conformational stability of ribonuclease T1 and mutant Glu-58----Ala. PMID:1980207
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 PDB Entries

Structure ID Release Date Resolution Structure Title
1B2M 1998-11-27T00:00:00+0000 2.0 THREE-DIMENSIONAL STRUCTURE OF RIBONULCEASE T1 COMPLEXED WITH AN ISOSTERIC PHOSPHONATE ANALOGUE OF GPU: ALTERNATE SUBSTRATE BINDING MODES AND CATALYSIS.
1BIR 1996-01-04T00:00:00+0000 1.8 RIBONUCLEASE T1, PHE 100 TO ALA MUTANT COMPLEXED WITH 2' GMP
1BU4 1998-09-11T00:00:00+0000 1.9 RIBONUCLEASE 1 COMPLEX WITH 2'GMP
1BVI 1998-09-15T00:00:00+0000 1.9 RIBONUCLEASE T1 (WILDTYPE) COMPLEXED WITH 2'GMP
1CH0 1999-03-30T00:00:00+0000 2.3 RNASE T1 VARIANT WITH ALTERED GUANINE BINDING SEGMENT
1DET 1996-02-20T00:00:00+0000 1.8 RIBONUCLEASE T1 CARBOXYMETHYLATED AT GLU 58 IN COMPLEX WITH 2'GMP
1FYS 2000-10-03T00:00:00+0000 2.0 Ribonuclease T1 V16C mutant
1FZU 2000-10-04T00:00:00+0000 1.8 RNAse T1 V78A mutant
1G02 2000-10-05T00:00:00+0000 1.86 Ribonuclease T1 V16S mutant
1GSP 1997-11-28T00:00:00+0000 2.2 RIBONUCLEASE T1 COMPLEXED WITH 2',3'-CGPS, 1 DAY

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
100.0 Guanyl-specific ribonuclease T1 P00651 RNT1_ASPOR