The structure of thioredoxin (Trx) from A. acidocaldarius (previously named Bacillus acidocaldarius) (BacTrx) and from Escherichia coli (EcoTrx) was studied by FTIR spectroscopy. Two mutants of BacTrx (K18G and R82E) were also analyzed. The data revealed similar secondary structures in all proteins, but BacTrx and its mutants showed a more compact structure than EcoTrx. In BacTrx and its mutants, the compactness was pD-dependent. All proteins revealed the existence of a molten globule (MG)-like state. At pD 5.8, the temp. at which this state was detected was higher in BacTrx and decreased in the different proteins in the following order: BacTrx > R82E > K18G > EcoTrx. At neutral or basic pD, the MG-like state was detected at the same temp. in both BacTrx and R82E, whereas it was found at the same temp. in all pD values tested for EcoTrx. The thermal stability of the proteins was in the following order at all pD values tested: BacTrx > R82E > K18G > EcoTrx, and was lower for each protein at pD 8.4 than at neutral or acidic pD values. The formation of protein aggregates, brought about by thermal denaturation, were obsd. for BacTrx and K18G at all pD values tested, whereas they were present in R82E and EcoTrx samples only at pD 5.8. The results indicated that a single mutation might affect the structural properties of a protein, including its propensity to aggregate at high temps. The data also indicated a possible application of FTIR spectroscopy for assessing MG-like states in small proteins.
Structural and thermal stability analysis of Escherichia coli and Alicyclobacillus acidocaldarius thioredoxin revealed a molten globule-like state in thermal denaturation pathway of the proteins: an infrared spectroscopic study.
Pedone E;Rossi M;
2003
Abstract
The structure of thioredoxin (Trx) from A. acidocaldarius (previously named Bacillus acidocaldarius) (BacTrx) and from Escherichia coli (EcoTrx) was studied by FTIR spectroscopy. Two mutants of BacTrx (K18G and R82E) were also analyzed. The data revealed similar secondary structures in all proteins, but BacTrx and its mutants showed a more compact structure than EcoTrx. In BacTrx and its mutants, the compactness was pD-dependent. All proteins revealed the existence of a molten globule (MG)-like state. At pD 5.8, the temp. at which this state was detected was higher in BacTrx and decreased in the different proteins in the following order: BacTrx > R82E > K18G > EcoTrx. At neutral or basic pD, the MG-like state was detected at the same temp. in both BacTrx and R82E, whereas it was found at the same temp. in all pD values tested for EcoTrx. The thermal stability of the proteins was in the following order at all pD values tested: BacTrx > R82E > K18G > EcoTrx, and was lower for each protein at pD 8.4 than at neutral or acidic pD values. The formation of protein aggregates, brought about by thermal denaturation, were obsd. for BacTrx and K18G at all pD values tested, whereas they were present in R82E and EcoTrx samples only at pD 5.8. The results indicated that a single mutation might affect the structural properties of a protein, including its propensity to aggregate at high temps. The data also indicated a possible application of FTIR spectroscopy for assessing MG-like states in small proteins.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
