title: Homo-dimeric recombinant dihydrofolate reductase from Thermotoga maritima shows extreme intrinsic stability.
abstract: Dihydrofolate reductase (DHFR) from the hyperthermophilic bacterium Thermotoga maritima was cloned and expressed in Escherichia coli. Sequence determination of the reported dyrA gene was repeated, and a corrected version deposited in the nucleotide sequence databank (accession number Y11021). Ultracentrifugational analysis and gel permeation chromatography prove that the enzyme forms a stable homodimer. The enzyme exhibits long-term stability at physiological temperature (80 degrees C) and in the presence of high denaturant concentrations (half-time in 6 M guanidinium chloride: 24h). Alignments of DHFRS from different species, as well as comparative modeling based on the homology to the crystal structures of the enzyme from prokaryotes and eukaryotes, were used to generate a model of the three-dimensional structure. The apoenzyme was crystallized and a data set was collected to a resolution of about 2 A.
authors.affiliation: Institut für Biophysik und Physikalische Biochemie, Universität Regensburg, Germany.
authors.forename: T
authors.initials: T
authors.lastname: Dams
authors.forename: G
authors.initials: G
authors.lastname: Böhm
authors.forename: G
authors.initials: G
authors.lastname: Auerbach
authors.forename: G
authors.initials: G
authors.lastname: Bader
authors.forename: H
authors.initials: H
authors.lastname: Schurig
authors.forename: R
authors.initials: R
authors.lastname: Jaenicke
mesh_terms: D000595:Amino Acid Sequence; D019281:Dimerization; D004795:Enzyme Stability; D006089:Gram-Negative Anaerobic Bacteria / Q000201:enzymology*; D006358:Hot Temperature; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013050:Spectrometry, Fluorescence; D013762:Tetrahydrofolate Dehydrogenase / Q000737:chemistry / Q000378:metabolism*
chemical_list: D011994:Recombinant Proteins; D013762:Tetrahydrofolate Dehydrogenase