1. Biochemical characterization of DNA-binding proteins from Pyrobaculum aerophilum and Aeropyrum pernix.
- Author
-
Hardy CD and Martin PK
- Subjects
- Aeropyrum genetics, Aeropyrum metabolism, Archaeal Proteins genetics, Archaeal Proteins metabolism, DNA, Archaeal genetics, DNA, Archaeal metabolism, DNA, Single-Stranded genetics, DNA, Single-Stranded metabolism, DNA-Binding Proteins genetics, DNA-Binding Proteins metabolism, Genome, Archaeal physiology, Kinetics, Open Reading Frames physiology, Protein Binding physiology, Pyrobaculum genetics, Pyrobaculum metabolism, Recombinant Proteins chemistry, Recombinant Proteins genetics, Recombinant Proteins metabolism, Aeropyrum chemistry, Archaeal Proteins chemistry, DNA, Archaeal chemistry, DNA, Single-Stranded chemistry, DNA-Binding Proteins chemistry, Pyrobaculum chemistry
- Abstract
Several representatives of the Crenarchaeal branch of the Archaea contain highly abundant, small, positively charged proteins exemplified by the Sso7d protein from Sulfolobus solfataricus. These proteins bind to DNA in a non-sequence-specific manner. Using publicly available genomic sequence information, we identified a second class of small Crenarchaeal DNA-binding proteins represented by the Pyrobaculum aerophilum open reading frame 3192-encoded (Pae3192) protein and its paralogs. We investigated the biochemical properties of the Pae3192 protein and an orthologous protein (Ape1322b) from Aeropyrum pernix in side-by-side experiments with the Sso7d protein. We demonstrate that the recombinant Ape1322b, Pae3192 and Sso7d proteins bind to DNA and that the DNA-protein complexes formed are slightly different for each protein. We show that like Sso7d, Pae3192 constrains negative supercoils in DNA. In addition, we show that all three proteins raise the melting temperature of duplex DNA upon binding. Finally, we present the equilibrium affinity constants and kinetic association constants of each protein for single-stranded and double-stranded DNA.
- Published
- 2008
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