Identification of a novel archaebacterial thioredoxin: determination of function through structure.
As part of a high-throughput, structural proteomic project we have used NMR spectroscopy to determine the solution structure and ascertain the function of a previously unknown, conserved protein (MtH895) from the thermophilic archeon Methanobacterium thermoautotrophicum. Our findings indicate that MtH895 contains a central four-stranded beta-sheet core surrounded by two helices on one side and a third on the other. It has an overall fold superficially similar to that of a glutaredoxin. However, detailed analysis of its three-dimensional structure along with molecular docking simulations of its interaction with T7 DNA polymerase (a thioredoxin-specific substrate) and comparisons with other known members of the thioredoxin/glutaredoxin family of proteins strongly suggest that MtH895 is more akin to a thioredoxin. Furthermore, measurement of the pK(a) values of its active site thiols along with direct measurements of the thioredoxin/glutaredoxin activity has confirmed that MtH895 is, indeed, a thioredoxin and exhibits no glutaredoxin activity. We have also identified a group of previously unknown proteins from several other archaebacteria that have significant (34-44%) sequence identity with MtH895. These proteins have unusual active site -CXXC- motifs not found in any known thioredoxin or glutaredoxin. On the basis of the results presented here, we predict that these small proteins are all members of a new class of truncated thioredoxins.
Bhattacharyya
Sudeepa
S
Faculty of Pharmacy and Pharmaceutical Sciences, University of Alberta, Edmonton, Alberta, Canada.
Habibi-Nazhad
Bahram
B
Amegbey
Godwin
G
Slupsky
Carolyn M
CM
Yee
Adelinda
A
Arrowsmith
Cheryl
C
Wishart
David S
DS
D000595:Amino Acid Sequence; D001665:Binding Sites; D017124:Conserved Sequence; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D017014:Methanobacterium / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D016415:Sequence Alignment; D012996:Solutions; D013816:Thermodynamics; D013879:Thioredoxins / Q000737:chemistry* / Q000378:metabolism*
D012996:Solutions; D013879:Thioredoxins; C034970:bacteriophage T7 induced DNA polymerase; D004259:DNA-Directed DNA Polymerase