Keywords: Deamidation, Folding, Isoaspartate, Mass Spectrometry, Protein Structure-Function, Ribonuclease A, Amino Acid Substitution, Article, Deamination, Disulfide Bond, Enzyme Analysis, Enzyme Structure, Priority Journal, Protein Folding, Structure Analysis, Animals, Cattle, Glutathione, Hydrogen Bonding, Kinetics, Oxidation-Reduction, Pancreatic,
Affiliations: Dipartimento di Chimica, Università degli Studi di Salerno, Salerno, Italy
Centro di Studio di Biocristallografia CNR, c/o Dipartimento di Chimica, Università degli Studi di Napoli Federico II, Italy
Centro Internazionale di Servizi Spettrometria di Massa, CNR-Università di Napoli, Italy
CEINGE, Biotecnologie Avanzate, Scrl, Napoli, Italy
References: Not available.
Effect of deamidation on folding of ribonuclease
The folding of ribonuclease A (RNase A) has been extensively studied by characterizing the disulfide containing intermediates using different experimental conditions and analytical techniques. So far, some aspects still remain unclear such as the role of the loop 65-72 in the folding pathway. We have studied the oxidative folding of a RNase A derivative containing at position 67 the substitution Asn --> isoAsp where the local structure of the loop 65-72 has been modified keeping intact the C65-C72 disulfide bond. By comparing the folding behavior of this mutant to that of the wild-type protein, we found that the deamidation significantly decreases the folding rate and alters the folding pathway of RNase A. Results presented here shed light on the role of the 65-72 region in the folding process of RNase A and also clarifies the effect of the deamidation on the folding/unfolding processes. On a more general ground, this study represents the first characterization of the intermediates produced along the folding of a deamidated protein.