Increase of RNase a N-terminus polarity or C-terminus apolarity changes the two domains' propensity to swap and form the two dimeric conformers of the protein.


Abstract

Do the polarities of the N-terminus or the apolarity of the C-terminus of bovine RNase A influence the relative yields of its two 3D domain-swapped dimeric conformers, the N-dimer and C-dimer? We have addressed this question by substituting Ala-4 or Ala-5 with serine (A4S and A5S mutants) or Ser-123 with alanine (S123A mutant) through site-directed mutagenesis. Both the polarity of the N-terminus and the apolarity of the C-terminus of RNase A were, therefore, increased. CD spectra revealed no significant differences between the secondary structures of the mutants and native RNase A. According to thermal denaturation analyses, the A4S and A5S mutants are less stable, and the S123A mutant is more stable than wild type RNase A. By subjecting the mutants under mild or drastic denaturing conditions, side-by-side with native and recombinant RNase A, to a thermally induced oligomerization procedure, the following results were obtained. (i) The N-terminal mutants showed a higher propensity, with respect to the native protein, to form N-dimers under mild unfolding conditions. (ii) The C-terminal mutant showed a higher propensity to form the C-dimer under severely unfolding conditions. These results are discussed in light of the relative stabilities of the various RNase A species under different environmental conditions, and we conclude that the hydrophilic or hydrophobic character of the RNase N-terminus or C-terminus can be an important variable governing the oligomerization of RNase A and possibly other proteins through the 3D domain-swapping mechanism. Study holds ProTherm entries: 20125, 20126, 20127, 20128, 20129, 20130, 20131, 20132, 20133, 20134, 20135, 20136, 20137, 20138, 20139, 20140, 20141, 20142, 20143, 20144, 20145 Extra Details: apolarity, site-directed mutagenesis, oligomerization, RNase A

Submission Details

ID: D3w22ayK

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:52 p.m.

Version: 1

Publication Details
Gotte G;Donadelli M;Laurents DV;Vottariello F;Morbio M;Libonati M,Biochemistry (2006) Increase of RNase a N-terminus polarity or C-terminus apolarity changes the two domains' propensity to swap and form the two dimeric conformers of the protein. PMID:16953565
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
6QMN 2019-02-07T00:00:00+0000 2.31 Crystal structure of a Ribonuclease A-Onconase chimera
1A2W 1998-01-12T00:00:00+0000 2.1 CRYSTAL STRUCTURE OF A 3D DOMAIN-SWAPPED DIMER OF BOVINE PANCREATIC RIBONUCLEASE A
1A5P 1998-02-17T00:00:00+0000 1.6 C[40,95]A VARIANT OF BOVINE PANCREATIC RIBONUCLEASE A
1A5Q 1998-02-17T00:00:00+0000 2.3 P93A VARIANT OF BOVINE PANCREATIC RIBONUCLEASE A
1AFK 1997-03-08T00:00:00+0000 1.7 CRYSTAL STRUCTURE OF RIBONUCLEASE A IN COMPLEX WITH 5'-DIPHOSPHOADENOSINE-3'-PHOSPHATE
1AFL 1997-03-08T00:00:00+0000 1.7 RIBONUCLEASE A IN COMPLEX WITH 5'-DIPHOSPHOADENOSINE 2'-PHOSPHATE AT 1.7 ANGSTROM RESOLUTION
1AFU 1997-03-14T00:00:00+0000 2.0 STRUCTURE OF RIBONUCLEASE A AT 2.0 ANGSTROMS FROM MONOCLINIC CRYSTALS
1AQP 1997-07-31T00:00:00+0000 2.0 RIBONUCLEASE A COPPER COMPLEX
1B6V 1999-01-18T00:00:00+0000 2.0 CRYSTAL STRUCTURE OF A HYBRID BETWEEN RIBONUCLEASE A AND BOVINE SEMINAL RIBONUCLEASE
1BEL 1995-12-21T00:00:00+0000 1.6 HYDROLASE PHOSPHORIC DIESTER, RNA

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
100.0 Ribonuclease pancreatic P61824 RNAS1_BISBI
100.0 Ribonuclease pancreatic P61823 RNAS1_BOVIN
96.8 Ribonuclease pancreatic P67926 RNAS1_CAPHI
96.8 Ribonuclease pancreatic P67927 RNAS1_SHEEP
95.2 Ribonuclease pancreatic P00657 RNAS1_BUBBU
96.0 Ribonuclease pancreatic P07847 RNAS1_AEPME
93.5 Ribonuclease pancreatic P07848 RNAS1_EUDTH
95.2 Ribonuclease pancreatic P00660 RNAS1_CONTA
92.7 Ribonuclease pancreatic P00668 RNAS1_ANTAM
90.3 Ribonuclease pancreatic P00662 RNAS1_GIRCA
96.0 Ribonuclease pancreatic Q29606 RNAS1_ORYLE