Conformation and conformational stability of ribonuclease A and its peptic derivative, des-(121-124)-ribonuclease

Biochemistry ◽  
1972 ◽  
Vol 11 (10) ◽  
pp. 1980-1990 ◽  
Author(s):  
David Puett ◽  
Betty K. Wasserman ◽  
Elaine Friebele
Keyword(s):  
Conformational Stability ◽  
Ribonuclease A

His···Asp Catalytic Dyad of Ribonuclease A:  Conformational Stability of the Wild-Type, D121N, D121A, and H119A Enzymes†

Biochemistry ◽  
1998 ◽  
Vol 37 (51) ◽  
pp. 17958-17964 ◽  
Author(s):  
David J. Quirk ◽  
Chiwook Park ◽  
James E. Thompson ◽  
Ronald T. Raines
Keyword(s):  
Conformational Stability ◽  
Ribonuclease A ◽  
Wild Type ◽  
Catalytic Dyad

scholarly journals Semisynthesis of Ribonuclease A using Intein-Mediated Protein Ligation

2002 ◽  
Vol 2 ◽  
pp. 1838-1842 ◽  
Author(s):  
Ulrich Arnold ◽  
Matthew P. Hinderaker ◽  
Ronald T. Raines
Keyword(s):  
Fusion Protein ◽  
Conformational Stability ◽  
Ribonuclease A ◽  
Rnase A ◽  
Amino Acid Residues ◽  
Wild Type ◽  
Thermal Stabilities ◽  
Protein Ligation ◽  
Hydrolysis Of ◽  
Control Protein

The introduction of non-natural amino acid residues or modules into proteins provides a new means to explore the basis for conformational stability, folding/unfolding behavior, or biological function. We exploited intein-mediated protein ligation to produce a semisynthetic ribonuclease A. Of the 124 residues of RNase A, residues 1–94 were linked to an intein. After expression of the fusion protein and thiol-induced cleavage, the RNase A(1–94) fragment possessed a C-terminal thioester. A peptide identical to the C-terminal residues 95–124 of RNase A (with residue 95 being cysteine) was successfully ligated to that thioester thereby reconstituting full-length wild-type RNase A. In mass spectrometry, this semisynthetic RNase A proved to be undistinguishable from the control protein, namely recombinant wild-type RNase A. Recombinant wild-type RNase A was obtained by expression of RNase A(1–124)–intein fusion protein followed by thiol-induced cleavage and hydrolysis of the thioester. Both proteins showed thermal stabilities (Tm) and catalytic activities comparable to the wild-type enzyme, indicating that both proteins folded properly. These results might serve as basis for the semisynthesis of RNase A variants containing non-natural modules in the aforementioned peptide.

scholarly journals Conformational Stability Is a Determinant of Ribonuclease A Cytotoxicity

2000 ◽  
Vol 275 (23) ◽  
pp. 17463-17467 ◽  
Author(s):  
Tony A. Klink ◽  
Ronald T. Raines
Keyword(s):  
Conformational Stability ◽  
Ribonuclease A

scholarly journals Contribution of disulfide bonds to the conformational stability and catalytic activity of ribonuclease A

2000 ◽  
Vol 267 (2) ◽  
pp. 566-572 ◽  
Author(s):  
Tony A. Klink ◽  
Kenneth J. Woycechowsky ◽  
Kimberly M. Taylor ◽  
Ronald T. Raines
Keyword(s):  
Catalytic Activity ◽  
Disulfide Bonds ◽  
Conformational Stability ◽  
Ribonuclease A
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