NMR solution structure of a novel thioredoxin from Bacillus acidocaldarius:
possible determinants of protein stability.

NMR solution structure of a novel thioredoxin from Bacillus acidocaldarius possible determinants of protein stability.

The thioredoxin (Trx) from Bacillus acidocaldarius (BacTrx) was purified to
homogeneity by anion-exchange chromatography and gel-filtration chromatography,
based on its ability to catalyse the dithiothreitol-dependent reduction of
bovine insulin disulphides. The protein has a molecular mass of 11577 Da,
determined by electrospray mass spectrometry, a pI of 4.2, and its primary
structure was obtained by automated Edman degradation after cleavage with
trypsin and cyanogen bromide. The sequences of known bacterial Trxs were
aligned at the active site: BacTrx has an identity ranging from 45 to 53% with
all sequences except that of the unusual Anabaena strain 7120 Trx (37%
identity). The gene coding for BacTrx was isolated by a strategy based on PCR
gene amplification and cloned in a plasmid downstream of a lac-derived promoter
sequence; the recombinant clone was used as the expression vector for
Escherichia coli. The expression was optimized by varying both the time of cell
growth and the time of exposure to the inducer isopropyl
beta-d-thiogalactoside; expressed BacTrx represents approx. 5% of the total
cytosolic protein. CD spectra and differential scanning calorimetry
measurements demonstrated that BacTrx is endowed with a higher conformational
heat stability than the Trx from E. coli. Nanogravimetry experiments showed a
lower content of bound water in BacTrx than in E. coli Trx, and a transition
temperature approx. 10 degrees C higher for BacTrx. The three-dimensional model
of the oxidized form of BacTrx was constructed by a comparative molecular
modelling technique, using E. coli Trx and Anabaena strain 7120 Trx as
reference proteins. Increased networks of ion-pairs and shorter loops emerged
as major features of the BacTrx structure compared with those of the template
proteins. The findings are discussed in the light of the current knowledge
about molecular determinants of protein stability.

Bartolucci et al. (1997)Biochem. J. 328, 277-285
Thioredoxin from Bacillus acidocaldarius: characterization, high-level expression in Escherichia coli and molecular modelling.

Thioredoxin from Bacillus acidocaldarius

The Crystal Structure Of The K18g Mutant Of The Thioredoxin From Alicyclobacillus Acidocaldarius

The Crystal Structure Of The Mutant R82e Of Thioredoxin From Alicyclobacillus Acidocaldarius

Solution Structure Of The Thioredoxin From Bacillus Acidocaldarius

Solution Structure Of The K18gR82E ALICYCLOBACILLUS Acidocaldarius Thioredoxin Mutant

thioredoxin [Alicyclobacillus acidocaldarius subsp. acidocaldarius DSM 446]

thioredoxin [Alicyclobacillus acidocaldarius subsp. acidocaldarius Tc-4-1]

thioredoxin [Alicyclobacillus acidocaldarius LAA1]

thioredoxin [Alicyclobacillus acidocaldarius]

Alicyclobacillus acidocaldarius subsp. acidocaldarius

SIMULATED ANNEALING, RESTRAINED MOLECULAR DYNAMICS AND RESTRAINED ENERGY